RESUMO
The choroid plexus (ChP) is a vital brain barrier and source of cerebrospinal fluid (CSF). Here, we use longitudinal two-photon imaging in awake mice and single-cell transcriptomics to elucidate the mechanisms of ChP regulation of brain inflammation. We used intracerebroventricular injections of lipopolysaccharides (LPS) to model meningitis in mice and observed that neutrophils and monocytes accumulated in the ChP stroma and surged across the epithelial barrier into the CSF. Bi-directional recruitment of monocytes from the periphery and, unexpectedly, macrophages from the CSF to the ChP helped eliminate neutrophils and repair the barrier. Transcriptomic analyses detailed the molecular steps accompanying this process and revealed that ChP epithelial cells transiently specialize to nurture immune cells, coordinating their recruitment, survival, and differentiation as well as regulation of the tight junctions that control the permeability of the ChP brain barrier. Collectively, we provide a mechanistic understanding and a comprehensive roadmap of neuroinflammation at the ChP brain barrier.
Assuntos
Barreira Hematoencefálica , Plexo Corióideo , Lipopolissacarídeos , Macrófagos , Doenças Neuroinflamatórias , Neutrófilos , Plexo Corióideo/metabolismo , Animais , Camundongos , Doenças Neuroinflamatórias/metabolismo , Barreira Hematoencefálica/metabolismo , Macrófagos/metabolismo , Macrófagos/imunologia , Neutrófilos/metabolismo , Neutrófilos/imunologia , Camundongos Endogâmicos C57BL , Monócitos/metabolismo , Masculino , Junções Íntimas/metabolismo , Células Epiteliais/metabolismo , FemininoRESUMO
Barrier tissue immune responses are regulated in part by nociceptors. Nociceptor ablation alters local immune responses at peripheral sites and within draining lymph nodes (LNs). The mechanisms and significance of nociceptor-dependent modulation of LN function are unknown. Using high-resolution imaging, viral tracing, single-cell transcriptomics, and optogenetics, we identified and functionally tested a sensory neuro-immune circuit that is responsive to lymph-borne inflammatory signals. Transcriptomics profiling revealed that multiple sensory neuron subsets, predominantly peptidergic nociceptors, innervate LNs, distinct from those innervating surrounding skin. To uncover LN-resident cells that may interact with LN-innervating sensory neurons, we generated a LN single-cell transcriptomics atlas and nominated nociceptor target populations and interaction modalities. Optogenetic stimulation of LN-innervating sensory fibers triggered rapid transcriptional changes in the predicted interacting cell types, particularly endothelium, stromal cells, and innate leukocytes. Thus, a unique population of sensory neurons monitors peripheral LNs and may locally regulate gene expression.
Assuntos
Imunomodulação , Linfonodos/imunologia , Linfonodos/inervação , Células Receptoras Sensoriais/imunologia , Potenciais de Ação , Animais , Inflamação/patologia , Camundongos , Nociceptores/metabolismo , Optogenética , Peptídeos/metabolismo , Pele/inervação , Sistema Nervoso Simpático/fisiologia , Receptores Toll-Like/agonistas , Receptores Toll-Like/metabolismoRESUMO
SARS-CoV-2 infection can cause severe respiratory COVID-19. However, many individuals present with isolated upper respiratory symptoms, suggesting potential to constrain viral pathology to the nasopharynx. Which cells SARS-CoV-2 primarily targets and how infection influences the respiratory epithelium remains incompletely understood. We performed scRNA-seq on nasopharyngeal swabs from 58 healthy and COVID-19 participants. During COVID-19, we observe expansion of secretory, loss of ciliated, and epithelial cell repopulation via deuterosomal cell expansion. In mild and moderate COVID-19, epithelial cells express anti-viral/interferon-responsive genes, while cells in severe COVID-19 have muted anti-viral responses despite equivalent viral loads. SARS-CoV-2 RNA+ host-target cells are highly heterogenous, including developing ciliated, interferon-responsive ciliated, AZGP1high goblet, and KRT13+ "hillock"-like cells, and we identify genes associated with susceptibility, resistance, or infection response. Our study defines protective and detrimental responses to SARS-CoV-2, the direct viral targets of infection, and suggests that failed nasal epithelial anti-viral immunity may underlie and precede severe COVID-19.
Assuntos
COVID-19/imunologia , COVID-19/virologia , Imunidade , SARS-CoV-2/fisiologia , Índice de Gravidade de Doença , Adulto , Idoso , Efeito Espectador , COVID-19/genética , Estudos de Coortes , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Nasofaringe/patologia , Nasofaringe/virologia , RNA Viral/análise , RNA Viral/genética , Mucosa Respiratória/patologia , Mucosa Respiratória/virologia , Transcrição Gênica , Carga ViralRESUMO
Genome-wide association studies (GWAS) have revealed risk alleles for ulcerative colitis (UC). To understand their cell type specificities and pathways of action, we generate an atlas of 366,650 cells from the colon mucosa of 18 UC patients and 12 healthy individuals, revealing 51 epithelial, stromal, and immune cell subsets, including BEST4+ enterocytes, microfold-like cells, and IL13RA2+IL11+ inflammatory fibroblasts, which we associate with resistance to anti-TNF treatment. Inflammatory fibroblasts, inflammatory monocytes, microfold-like cells, and T cells that co-express CD8 and IL-17 expand with disease, forming intercellular interaction hubs. Many UC risk genes are cell type specific and co-regulated within relatively few gene modules, suggesting convergence onto limited sets of cell types and pathways. Using this observation, we nominate and infer functions for specific risk genes across GWAS loci. Our work provides a framework for interrogating complex human diseases and mapping risk variants to cell types and pathways.
Assuntos
Colite Ulcerativa/patologia , Colo/metabolismo , Adulto , Idoso , Anticorpos Monoclonais/uso terapêutico , Bestrofinas/metabolismo , Antígenos CD8/metabolismo , Estudos de Casos e Controles , Colite Ulcerativa/tratamento farmacológico , Colite Ulcerativa/metabolismo , Colo/patologia , Enterócitos/citologia , Enterócitos/metabolismo , Feminino , Loci Gênicos , Estudo de Associação Genômica Ampla , Humanos , Interleucina-17/metabolismo , Masculino , Pessoa de Meia-Idade , Fatores de Risco , Linfócitos T/citologia , Linfócitos T/metabolismo , Trombospondinas/metabolismo , Fator de Necrose Tumoral alfa/imunologia , Fator de Necrose Tumoral alfa/metabolismo , Adulto JovemRESUMO
The nasal mucosa is often the initial site of respiratory viral infection, replication, and transmission. Understanding how infection shapes tissue-scale primary and memory responses is critical for designing mucosal therapeutics and vaccines. We generated a single-cell RNA-sequencing atlas of the murine nasal mucosa, sampling three regions during primary influenza infection and rechallenge. Compositional analysis revealed restricted infection to the respiratory mucosa with stepwise changes in immune and epithelial cell subsets and states. We identified and characterized a rare subset of Krt13+ nasal immune-interacting floor epithelial (KNIIFE) cells, which concurrently increased with tissue-resident memory T (TRM)-like cells. Proportionality analysis, cell-cell communication inference, and microscopy underscored the CXCL16-CXCR6 axis between KNIIFE and TRM cells. Secondary influenza challenge induced accelerated and coordinated myeloid and lymphoid responses without epithelial proliferation. Together, this atlas serves as a reference for viral infection in the upper respiratory tract and highlights the efficacy of local coordinated memory responses.
Assuntos
Memória Imunológica , Células T de Memória , Mucosa Nasal , Infecções por Orthomyxoviridae , Animais , Memória Imunológica/imunologia , Camundongos , Mucosa Nasal/virologia , Mucosa Nasal/imunologia , Infecções por Orthomyxoviridae/imunologia , Infecções por Orthomyxoviridae/virologia , Células T de Memória/imunologia , Células Epiteliais/imunologia , Células Epiteliais/virologia , Camundongos Endogâmicos C57BL , Humanos , Análise de Célula Única , Influenza Humana/imunologia , Influenza Humana/virologia , Feminino , Receptores CXCR6/metabolismo , Receptores CXCR6/imunologia , Vírus da Influenza A/imunologia , Vírus da Influenza A/fisiologiaRESUMO
In the small intestine, a niche of accessory cell types supports the generation of mature epithelial cell types from intestinal stem cells (ISCs). It is unclear, however, if and how immune cells in the niche affect ISC fate or the balance between self-renewal and differentiation. Here, we use single-cell RNA sequencing (scRNA-seq) to identify MHC class II (MHCII) machinery enrichment in two subsets of Lgr5+ ISCs. We show that MHCII+ Lgr5+ ISCs are non-conventional antigen-presenting cells in co-cultures with CD4+ T helper (Th) cells. Stimulation of intestinal organoids with key Th cytokines affects Lgr5+ ISC renewal and differentiation in opposing ways: pro-inflammatory signals promote differentiation, while regulatory cells and cytokines reduce it. In vivo genetic perturbation of Th cells or MHCII expression on Lgr5+ ISCs impacts epithelial cell differentiation and IEC fate during infection. These interactions between Th cells and Lgr5+ ISCs, thus, orchestrate tissue-wide responses to external signals.
Assuntos
Diferenciação Celular , Autorrenovação Celular , Interleucina-10/metabolismo , Células-Tronco/citologia , Linfócitos T Auxiliares-Indutores/metabolismo , Animais , Diferenciação Celular/efeitos dos fármacos , Autorrenovação Celular/efeitos dos fármacos , Citocinas/farmacologia , Células Epiteliais/citologia , Células Epiteliais/metabolismo , Feminino , Antígenos de Histocompatibilidade Classe II/metabolismo , Sistema Imunitário/metabolismo , Intestinos/citologia , Intestinos/microbiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Organoides/citologia , Organoides/efeitos dos fármacos , Organoides/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Salmonella enterica/patogenicidade , Células-Tronco/metabolismo , Linfócitos T Auxiliares-Indutores/citologiaRESUMO
Technical, analytical, and ethical challenges have obscured our understanding of immune cell subset ontogeny during human fetal development. Recently published in Science, Suo et al. (2022) apply multiple single-cell and spatial tools to provide a comprehensive roadmap during human gestation.
Assuntos
Desenvolvimento Embrionário , Desenvolvimento Fetal , Feminino , Feto , Humanos , GravidezRESUMO
Mycobacterium tuberculosis lung infection results in a complex multicellular structure: the granuloma. In some granulomas, immune activity promotes bacterial clearance, but in others, bacteria persist and grow. We identified correlates of bacterial control in cynomolgus macaque lung granulomas by co-registering longitudinal positron emission tomography and computed tomography imaging, single-cell RNA sequencing, and measures of bacterial clearance. Bacterial persistence occurred in granulomas enriched for mast, endothelial, fibroblast, and plasma cells, signaling amongst themselves via type 2 immunity and wound-healing pathways. Granulomas that drove bacterial control were characterized by cellular ecosystems enriched for type 1-type 17, stem-like, and cytotoxic T cells engaged in pro-inflammatory signaling networks involving diverse cell populations. Granulomas that arose later in infection displayed functional characteristics of restrictive granulomas and were more capable of killing Mtb. Our results define the complex multicellular ecosystems underlying (lack of) granuloma resolution and highlight host immune targets that can be leveraged to develop new vaccine and therapeutic strategies for TB.
Assuntos
Mycobacterium tuberculosis , Fibrose Pulmonar , Tuberculose , Animais , Ecossistema , Granuloma , Pulmão , Macaca fascicularis , Fibrose Pulmonar/patologiaRESUMO
For new principal investigators, the first years are key to getting a laboratory off the ground and running. COVID-19 has changed the world, bringing on unforeseen difficulties and challenges at every level. We asked these investigators to share their experiences in navigating the unique environment since the start of the pandemic-what has changed in their vision for their laboratory, how they have adapted, and what advice they can share with others in a similar situation.
Assuntos
COVID-19/epidemiologia , Laboratórios , Adaptação Psicológica , Pesquisa Biomédica/tendências , COVID-19/psicologia , Comunicação , Humanos , Laboratórios/tendências , Pessoal de Laboratório/psicologia , Pessoal de Laboratório/tendências , SARS-CoV-2RESUMO
High-throughput single-cell RNA-sequencing (scRNA-seq) methodologies enable characterization of complex biological samples by increasing the number of cells that can be profiled contemporaneously. Nevertheless, these approaches recover less information per cell than low-throughput strategies. To accurately report the expression of key phenotypic features of cells, scRNA-seq platforms are needed that are both high fidelity and high throughput. To address this need, we created Seq-Well S3 ("Second-Strand Synthesis"), a massively parallel scRNA-seq protocol that uses a randomly primed second-strand synthesis to recover complementary DNA (cDNA) molecules that were successfully reverse transcribed but to which a second oligonucleotide handle, necessary for subsequent whole transcriptome amplification, was not appended due to inefficient template switching. Seq-Well S3 increased the efficiency of transcript capture and gene detection compared with that of previous iterations by up to 10- and 5-fold, respectively. We used Seq-Well S3 to chart the transcriptional landscape of five human inflammatory skin diseases, thus providing a resource for the further study of human skin inflammation.
Assuntos
Sequenciamento de Nucleotídeos em Larga Escala/métodos , Inflamação/genética , RNA Citoplasmático Pequeno/genética , Pele/patologia , Animais , Linhagem Celular , DNA Complementar/genética , Células HEK293 , Humanos , Camundongos , Células NIH 3T3 , Análise de Sequência de RNA/métodos , Análise de Célula Única/métodos , Transcrição Gênica/genética , Transcriptoma/genéticaRESUMO
Tissue ecosystems are cellular communities that maintain setpoints through a network of intercellular interactions. We position health and chronic inflammatory disease as alternative stable setpoints that are: (1) robust to perturbation, and (2) capable of adaptation and memory. Inflammatory memory-the storage of prior experience to durably influence future responsiveness-is central to how tissue ecosystems may be pushed past tipping points that stabilize disease over health. Here, we develop a reductionist framework of circuit motifs that recur in tissue setpoints. In type 2 immunity, we distinctly find the emergence of two-cell positive feedback motifs. By contrast, directional motif relays and three-cell networks feature more prominently in type 1 and 17 responses. We propose that these differences guide the ecological networks established after surpassing tipping points and associate closely with therapeutic responsiveness. We highlight opportunities to improve our current knowledge of how circuit motifs interact when building towards tissue-level networks across adaptation and memory. By developing new tools for circuit motif nomination and applying them to temporal profiling of tissue ecosystems, we hope to dissect the stability of the chronic inflammatory setpoint and open therapeutic avenues for rewriting memory to restore health.
RESUMO
Barrier tissue dysfunction is a fundamental feature of chronic human inflammatory diseases1. Specialized subsets of epithelial cells-including secretory and ciliated cells-differentiate from basal stem cells to collectively protect the upper airway2-4. Allergic inflammation can develop from persistent activation5 of type 2 immunity6 in the upper airway, resulting in chronic rhinosinusitis, which ranges in severity from rhinitis to severe nasal polyps7. Basal cell hyperplasia is a hallmark of severe disease7-9, but it is not known how these progenitor cells2,10,11 contribute to clinical presentation and barrier tissue dysfunction in humans. Here we profile primary human surgical chronic rhinosinusitis samples (18,036 cells, n = 12) that span the disease spectrum using Seq-Well for massively parallel single-cell RNA sequencing12, report transcriptomes for human respiratory epithelial, immune and stromal cell types and subsets from a type 2 inflammatory disease, and map key mediators. By comparison with nasal scrapings (18,704 cells, n = 9), we define signatures of core, healthy, inflamed and polyp secretory cells. We reveal marked differences between the epithelial compartments of the non-polyp and polyp cellular ecosystems, identifying and validating a global reduction in cellular diversity of polyps characterized by basal cell hyperplasia, concomitant decreases in glandular cells, and phenotypic shifts in secretory cell antimicrobial expression. We detect an aberrant basal progenitor differentiation trajectory in polyps, and propose cell-intrinsic13, epigenetic14,15 and extrinsic factors11,16,17 that lock polyp basal cells into this uncommitted state. Finally, we functionally demonstrate that ex vivo cultured basal cells retain intrinsic memory of IL-4/IL-13 exposure, and test the potential for clinical blockade of the IL-4 receptor α-subunit to modify basal and secretory cell states in vivo. Overall, we find that reduced epithelial diversity stemming from functional shifts in basal cells is a key characteristic of type 2 immune-mediated barrier tissue dysfunction. Our results demonstrate that epithelial stem cells may contribute to the persistence of human disease by serving as repositories for allergic memories.
Assuntos
Hipersensibilidade/imunologia , Hipersensibilidade/patologia , Mucosa Respiratória/imunologia , Mucosa Respiratória/patologia , Células-Tronco/imunologia , Células-Tronco/patologia , Adolescente , Adulto , Idoso , Estudos de Casos e Controles , Células Cultivadas , Epigênese Genética , Células Epiteliais/imunologia , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Humanos , Hiperplasia/metabolismo , Hiperplasia/patologia , Interleucina-13/imunologia , Interleucina-4/imunologia , Subunidade alfa de Receptor de Interleucina-4/antagonistas & inibidores , Subunidade alfa de Receptor de Interleucina-4/imunologia , Pessoa de Meia-Idade , Pólipos Nasais/imunologia , Pólipos Nasais/patologia , Rinite/imunologia , Rinite/patologia , Análise de Sequência de RNA , Análise de Célula Única , Sinusite/imunologia , Sinusite/patologia , Transcrição Gênica , Transcriptoma , Adulto JovemRESUMO
Standalone and consortia-led single-cell atlases of healthy and diseased human airways generated with single-cell RNA-sequencing (scRNA-seq) have ushered in a new era in respiratory research. Numerous discoveries, including the pulmonary ionocyte, potentially novel cell fates, and a diversity of cell states among common and rare epithelial cell types have highlighted the extent of cellular heterogeneity and plasticity in the respiratory tract. scRNA-seq has also played a pivotal role in our understanding of host-virus interactions in coronavirus disease 2019 (COVID-19). However, as our ability to generate large quantities of scRNA-seq data increases, along with a growing number of scRNA-seq protocols and data analysis methods, new challenges related to the contextualisation and downstream applications of insights are arising. Here, we review the fundamental concept of cellular identity from the perspective of single-cell transcriptomics in the respiratory context, drawing attention to the need to generate reference annotations and to standardise the terminology used in literature. Findings about airway epithelial cell types, states and fates obtained from scRNA-seq experiments are compared and contrasted with information accumulated through the use of conventional methods. This review attempts to discuss major opportunities and to outline some of the key limitations of the modern-day scRNA-seq that need to be addressed to enable efficient and meaningful integration of scRNA-seq data from different platforms and studies, with each other as well as with data from other high-throughput sequencing-based genomic, transcriptomic and epigenetic analyses.
Assuntos
COVID-19 , Análise de Célula Única , Humanos , Análise de Sequência de RNA/métodos , Análise de Célula Única/métodos , COVID-19/genética , Perfilação da Expressão Gênica/métodos , Células Epiteliais , RNA/genéticaRESUMO
Cholesterol biosynthetic intermediates, such as lanosterol and desmosterol, are emergent immune regulators of macrophages in response to inflammatory stimuli or lipid overloading, respectively. However, the participation of these sterols in regulating macrophage functions in the physiological context of atherosclerosis, an inflammatory disease driven by the accumulation of cholesterol-laden macrophages in the artery wall, has remained elusive. Here, we report that desmosterol, the most abundant cholesterol biosynthetic intermediate in human coronary artery lesions, plays an essential role during atherogenesis, serving as a key molecule integrating cholesterol homeostasis and immune responses in macrophages. Depletion of desmosterol in myeloid cells by overexpression of 3ß-hydroxysterol Δ24-reductase (DHCR24), the enzyme that catalyzes conversion of desmosterol to cholesterol, promotes the progression of atherosclerosis. Single-cell transcriptomics in isolated CD45+CD11b+ cells from atherosclerotic plaques demonstrate that depletion of desmosterol increases interferon responses and attenuates the expression of antiinflammatory macrophage markers. Lipidomic and transcriptomic analysis of in vivo macrophage foam cells demonstrate that desmosterol is a major endogenous liver X receptor (LXR) ligand involved in LXR/retinoid X receptor (RXR) activation and thus macrophage foam cell formation. Decreased desmosterol accumulation in mitochondria promotes macrophage mitochondrial reactive oxygen species production and NLR family pyrin domain containing 3 (NLRP3)-dependent inflammasome activation. Deficiency of NLRP3 or apoptosis-associated speck-like protein containing a CARD (ASC) rescues the increased inflammasome activity and atherogenesis observed in desmosterol-depleted macrophages. Altogether, these findings underscore the critical function of desmosterol in the atherosclerotic plaque to dampen inflammation by integrating with macrophage cholesterol metabolism and inflammatory activation and protecting from disease progression.
Assuntos
Aterosclerose/tratamento farmacológico , Desmosterol/farmacologia , Inflamassomos/metabolismo , Inflamação/tratamento farmacológico , Ativação de Macrófagos/efeitos dos fármacos , Animais , Aterosclerose/metabolismo , Aterosclerose/patologia , Colesterol/metabolismo , Vasos Coronários , Células Espumosas/metabolismo , Humanos , Inflamação/metabolismo , Metabolismo dos Lipídeos , Receptores X do Fígado/metabolismo , Macrófagos/metabolismo , Masculino , Camundongos , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Oxirredutases atuantes sobre Doadores de Grupo CH-CH/genética , Oxirredutases atuantes sobre Doadores de Grupo CH-CH/metabolismo , Placa Aterosclerótica/metabolismo , Esteróis/metabolismoRESUMO
BACKGROUND: Dupilumab, a mAb targeting IL-4Rα, improves upper and lower airway symptoms in patients with aspirin-exacerbated respiratory disease (AERD), but the mechanisms leading to clinical improvement are not fully elucidated. OBJECTIVE: Our aim was to identify the mechanistic basis of clinical improvement in patients with AERD treated with dupilumab. METHODS: A total of 22 patients with AERD were treated with dupilumab for 3 months for severe asthma and/or chronic rhinosinusitis with nasal polyps. Clinical outcomes were assessed at baseline and at 1 and 3 months after initiation of dupilumab. Nasal fluid, urine, blood, and inferior turbinate scrapings were collected at the 3 time points for determination of mediator levels, cellular assays, and RNA sequencing. RESULTS: Participants had rapid improvement in clinical measures, including sense of smell, sinonasal symptoms, and lung function after 1 month of treatment with dupilumab; the improvements were sustained after 3 months of dupilumab. Baseline severity of smell loss was correlated with lower nasal prostaglandin E2 levels. Dupilumab increased nasal prostaglandin E2 level and decreased levels of nasal albumin, nasal and urinary leukotriene E4, and serum and nasal IgE. Transcripts related to epithelial dysfunction and leukocyte activation and migration were downregulated in inferior turbinate tissue after treatment with dupilumab. There were no dupilumab-induced changes in nasal eosinophilia. CONCLUSION: Inhibition of IL-4Rα in AERD led to rapid improvement in respiratory symptoms and smell, with a concomitant improvement in epithelial barrier function, a decrease in inflammatory eicosanoid levels, and an increase in the anti-inflammatory eicosanoid prostaglandin E2 level. The therapeutic effects of dupilumab are likely due to decreased IL-4Rα signaling on respiratory tissue granulocytes, epithelial cells, and B cells.
Assuntos
Asma Induzida por Aspirina , Pólipos Nasais , Rinite , Sinusite , Aspirina/efeitos adversos , Asma Induzida por Aspirina/diagnóstico , Doença Crônica , Eicosanoides , Humanos , Pólipos Nasais/induzido quimicamente , Pólipos Nasais/tratamento farmacológico , Prostaglandinas , Rinite/induzido quimicamente , Rinite/tratamento farmacológico , Sinusite/induzido quimicamente , Sinusite/tratamento farmacológicoRESUMO
BACKGROUND: Eosinophilic asthma and nasal polyposis are hallmarks of aspirin-exacerbated respiratory disease (AERD), and IL-5 inhibition has been shown to provide therapeutic benefit. However, IL-5Rα is expressed on many cells in addition to eosinophils, and the mechanisms by which IL-5 inhibition leads to clinical benefit in eosinophilic asthma and nasal polyposis are unlikely to be due exclusively to antieosinophil effects. OBJECTIVE: We sought to identify the mechanisms by which anti-IL-5 treatment with mepolizumab improves respiratory inflammation in AERD. METHODS: The clinical characteristics, circulating granulocytes, nasal scraping transcripts, eosinophilic cationic protein, tryptase, and antibody levels, and urinary and nasal eicosanoid levels were measured for 18 subjects with AERD who were taking mepolizumab and compared with those of 18 matched subjects with AERD who were not taking mepolizumab. RESULTS: Subjects taking mepolizumab had significantly fewer peripheral blood eosinophils and basophils, and those cells that remained had higher surface CRTH2 expression than did the cells from subjects not taking mepolizumab. Nasal prostaglandin F2α, prostaglandin D2 metabolites, leukotriene B4, and thromboxane levels were lower in subjects taking mepolizumab, as were urinary levels of tetranor-prostaglandin D2 and leukotriene E4. The nasal epithelial cell transcripts that were overexpressed among subjects with AERD who were taking mepolizumab were enriched for genes involved in tight junction formation and cilium organization. Nasal and urinary prostaglandin E2, tryptase, and antibody levels were not different between the 2 groups. CONCLUSION: IL-5 inhibition in AERD decreases production of inflammatory eicosanoids and upregulates tight junction-associated nasal epithelial cell transcripts, likely due to decreased IL-5 signaling on tissue mast cells, eosinophils, and epithelial cells. These direct effects on multiple relevant immune cells contribute to the mechanism of benefit afforded by mepolizumab.
Assuntos
Anticorpos Monoclonais Humanizados/administração & dosagem , Asma Induzida por Aspirina , Basófilos , Eosinófilos , Pólipos Nasais , Adolescente , Adulto , Idoso , Asma Induzida por Aspirina/tratamento farmacológico , Asma Induzida por Aspirina/imunologia , Asma Induzida por Aspirina/urina , Basófilos/imunologia , Basófilos/patologia , Eicosanoides/imunologia , Eicosanoides/urina , Eosinófilos/imunologia , Eosinófilos/patologia , Feminino , Humanos , Interleucina-5/imunologia , Subunidade alfa de Receptor de Interleucina-5/imunologia , Masculino , Pessoa de Meia-Idade , Pólipos Nasais/tratamento farmacológico , Pólipos Nasais/imunologia , Pólipos Nasais/urinaRESUMO
BACKGROUND & AIMS: Studies are needed to determine the mechanisms of mucosal dysregulation in patients with inflammatory bowel diseases (IBDs) and differences in inflammatory responses of patients with ulcerative colitis (UC) vs Crohn's disease (CD). We used mass cytometry (CyTOF) to characterize and compare immune cell populations in the mucosa and blood from patients with IBD and without IBD (controls) at single-cell resolution. METHODS: We performed CyTOF analysis of colonic mucosa samples (n = 87) and peripheral blood mononuclear cells (n = 85) from patients with active or inactive UC or CD and controls. We also performed single-cell RNA sequencing, flow cytometry, and RNA in situ hybridization analyses to validate key findings. We used random forest modeling to identify differences in signatures across subject groups. RESULTS: Compared with controls, colonic mucosa samples from patients with IBD had increased abundances of HLA-DR+CD38+ T cells, including T-regulatory cells that produce inflammatory cytokines; CXCR3+ plasmablasts; and IL1B+ macrophages and monocytes. Colonic mucosa samples from patients with UC were characterized by expansion of IL17A+ CD161+ effector memory T cells and IL17A+ T-regulatory cells; expansion of HLA-DR+CD56+ granulocytes; and reductions in type 3 innate lymphoid cells. Mucosal samples from patients with active CD were characterized by IL1B+HLA-DR+CD38+ T cells, IL1B+TNF+IFNG+ naïve B cells, IL1B+ dendritic cells (DCs), and IL1B+ plasmacytoid DCs. Peripheral blood mononuclear cells from patients with active CD differed from those of active UC in that the peripheral blood mononuclear cells from patients with CD had increased IL1B+ T-regulatory cells, IL1B+ DCs and IL1B+ plasmacytoid DCs, IL1B+ monocytes, and fewer group 1 innate lymphoid cells. Random forest modeling differentiated active UC from active CD in colonic mucosa and blood samples; top discriminating features included many of the cellular populations identified above. CONCLUSIONS: We used single-cell technologies to identify immune cell populations specific to mucosa and blood samples from patients with active or inactive CD and UC and controls. This information might be used to develop therapies that target specific cell populations in patients with different types of IBD.
Assuntos
Colite Ulcerativa/imunologia , Doença de Crohn/imunologia , Imunidade Celular , Imunofenotipagem/métodos , Adolescente , Adulto , Estudos de Casos e Controles , Criança , Colite Ulcerativa/sangue , Colite Ulcerativa/patologia , Colo/imunologia , Colo/patologia , Doença de Crohn/sangue , Doença de Crohn/patologia , Feminino , Citometria de Fluxo , Humanos , Mucosa Intestinal/imunologia , Mucosa Intestinal/patologia , Masculino , RNA-Seq , Análise de Célula Única , Adulto JovemRESUMO
BACKGROUND: The cause of severe nasal polyposis in aspirin-exacerbated respiratory disease (AERD) is unknown. Elevated antibody levels have been associated with disease severity in nasal polyps, but upstream drivers of local antibody production in nasal polyps are undetermined. OBJECTIVE: We sought to identify upstream drivers and phenotypic properties of local antibody-expressing cells in nasal polyps from subjects with AERD. METHODS: Sinus tissue was obtained from subjects with AERD, chronic rhinosinusitis (CRS) with nasal polyps (CRSwNP), CRS without nasal polyps, and controls without CRS. Tissue antibody levels were quantified via ELISA and immunohistochemistry and were correlated with disease severity. Antibody-expressing cells were profiled with single-cell RNA sequencing, flow cytometry, and immunofluorescence, with IL-5Rα function determined through IL-5 stimulation and subsequent RNA sequencing and quantitative PCR. RESULTS: Tissue IgE and IgG4 levels were elevated in AERD compared with in controls (P < .01 for IgE and P < .001 for IgG4 vs CRSwNP). Subjects with AERD whose nasal polyps recurred rapidly had higher IgE levels than did subjects with AERD, with slower regrowth (P = .005). Single-cell RNA sequencing revealed increased IL5RA, IGHG4, and IGHE in antibody-expressing cells from patients with AERD compared with antibody-expressing cells from patients with CRSwNP. There were more IL-5Rα+ plasma cells in the polyp tissue from those with AERD than in polyp tissue from those with CRSwNP (P = .026). IL-5 stimulation of plasma cells in vitro induced changes in a distinct set of transcripts. CONCLUSIONS: Our study identifies an increase in antibody-expressing cells in AERD defined by transcript enrichment of IL5RA and IGHG4 or IGHE, with confirmed surface expression of IL-5Rα and functional IL-5 signaling. Tissue IgE and IgG4 levels are elevated in AERD, and higher IgE levels are associated with faster nasal polyp regrowth. Our findings suggest a role for IL-5Rα+ antibody-expressing cells in facilitating local antibody production and severe nasal polyps in AERD.
Assuntos
Aspirina/efeitos adversos , Imunoglobulina E/metabolismo , Imunoglobulina G/metabolismo , Subunidade alfa de Receptor de Interleucina-5/metabolismo , Pólipos Nasais/metabolismo , Sinusite/metabolismo , Adulto , Idoso , Anticorpos/metabolismo , Feminino , Humanos , Interleucina-5/metabolismo , Masculino , Pessoa de Meia-Idade , Pólipos Nasais/induzido quimicamente , Plasmócitos/efeitos dos fármacos , Plasmócitos/metabolismo , Análise de Sequência de RNA/métodos , Sinusite/induzido quimicamente , Adulto JovemRESUMO
The skin has a dual function as a barrier and a sensory interface between the body and the environment. To protect against invading pathogens, the skin harbours specialized immune cells, including dermal dendritic cells (DDCs) and interleukin (IL)-17-producing γδ T (γδT17) cells, the aberrant activation of which by IL-23 can provoke psoriasis-like inflammation. The skin is also innervated by a meshwork of peripheral nerves consisting of relatively sparse autonomic and abundant sensory fibres. Interactions between the autonomic nervous system and immune cells in lymphoid organs are known to contribute to systemic immunity, but how peripheral nerves regulate cutaneous immune responses remains unclear. We exposed the skin of mice to imiquimod, which induces IL-23-dependent psoriasis-like inflammation. Here we show that a subset of sensory neurons expressing the ion channels TRPV1 and Nav1.8 is essential to drive this inflammatory response. Imaging of intact skin revealed that a large fraction of DDCs, the principal source of IL-23, is in close contact with these nociceptors. Upon selective pharmacological or genetic ablation of nociceptors, DDCs failed to produce IL-23 in imiquimod-exposed skin. Consequently, the local production of IL-23-dependent inflammatory cytokines by dermal γδT17 cells and the subsequent recruitment of inflammatory cells to the skin were markedly reduced. Intradermal injection of IL-23 bypassed the requirement for nociceptor communication with DDCs and restored the inflammatory response. These findings indicate that TRPV1(+)Nav1.8(+) nociceptors, by interacting with DDCs, regulate the IL-23/IL-17 pathway and control cutaneous immune responses.
Assuntos
Interleucina-23/imunologia , Nociceptores/metabolismo , Psoríase/imunologia , Psoríase/patologia , Células Receptoras Sensoriais/metabolismo , Pele/inervação , Pele/patologia , Aminoquinolinas , Animais , Modelos Animais de Doenças , Feminino , Imiquimode , Inflamação/induzido quimicamente , Inflamação/imunologia , Inflamação/patologia , Interleucina-17/biossíntese , Interleucina-17/imunologia , Interleucina-23/biossíntese , Interleucinas/biossíntese , Interleucinas/imunologia , Células de Langerhans/imunologia , Células de Langerhans/metabolismo , Linfonodos/imunologia , Linfonodos/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Canal de Sódio Disparado por Voltagem NAV1.8/metabolismo , Nociceptores/efeitos dos fármacos , Psoríase/induzido quimicamente , Células Receptoras Sensoriais/efeitos dos fármacos , Pele/citologia , Pele/imunologia , Linfócitos T/imunologia , Linfócitos T/metabolismo , Canais de Cátion TRPV/metabolismo , Interleucina 22RESUMO
Journal-based science communication is not accessible or comprehensible to a general public curious about science and eager for the next wave of scientific innovation. We propose an alternative medium for scientists to communicate their work to the general public in an engaging and digestible way through the use of whiteboard videos. We describe the process of producing science whiteboard videos and the benefits and challenges therein.