RESUMEN
The miR-15/16 family targets a large network of genes in T cells to restrict their cell cycle, memory formation, and survival. Upon T cell activation, miR-15/16 are downregulated, allowing rapid expansion of differentiated effector T cells to mediate a sustained response. Here, we used conditional deletion of miR-15/16 in regulatory T cells (Tregs) to identify immune functions of the miR-15/16 family in T cells. miR-15/16 are indispensable to maintain peripheral tolerance by securing efficient suppression by a limited number of Tregs. miR-15/16 deficiency alters expression of critical Treg proteins and results in accumulation of functionally impaired FOXP3loCD25loCD127hi Tregs. Excessive proliferation in the absence of miR-15/16 shifts Treg fate and produces an effector Treg phenotype. These Tregs fail to control immune activation, leading to spontaneous multi-organ inflammation and increased allergic inflammation in a mouse model of asthma. Together, our results demonstrate that miR-15/16 expression in Tregs is essential to maintain immune tolerance.
Asunto(s)
MicroARNs , Linfocitos T Reguladores , Animales , Ratones , MicroARNs/genética , MicroARNs/metabolismo , División Celular , Fenotipo , Inflamación/genética , Inflamación/metabolismo , Factores de Transcripción Forkhead/metabolismoRESUMEN
Despite robust literature associating IL-31 with pruritic inflammatory skin diseases, its influence on cutaneous inflammation and the interplay between inflammatory and neurosensory pathways remain unmapped. Here, we examined the consequences of disrupting Il31 and its receptor Il31ra in a mouse model of house dust mite (HDM)-induced allergic dermatitis. Il31-deficient mice displayed a deficit in HDM dermatitis-associated scratching, consistent with its well-established role as a pruritogen. In contrast, Il31 deficiency increased the number and proportion of cutaneous type 2 cytokine-producing CD4+ T cells and serum IgE in response to HDM. Furthermore, Il4ra+ monocytes and macrophages capable of fueling a feedforward type 2 inflammatory loop were selectively enriched in Il31ra-deficient HDM dermatitis skin. Thus, IL-31 is not strictly a proinflammatory cytokine but rather an immunoregulatory factor that limits the magnitude of type 2 inflammatory responses in skin. Our data support a model wherein IL-31 activation of IL31RA+ pruritoceptors triggers release of calcitonin gene-related protein (CGRP), which can mediate neurogenic inflammation, inhibit CD4+ T cell proliferation, and reduce T cell production of the type 2 cytokine IL-13. Together, these results illustrate a previously unrecognized neuroimmune pathway that constrains type 2 tissue inflammation in the setting of chronic cutaneous allergen exposure and may explain paradoxical dermatitis flares in atopic patients treated with anti-IL31RA therapy.
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Dermatitis Atópica , Inflamación Neurogénica , Animales , Ratones , Citocinas , Inmunidad , Pyroglyphidae , Piel/inmunología , Interleucinas/inmunología , Interleucinas/metabolismoRESUMEN
The miR-15/16 family is a highly expressed group of tumor suppressor miRNAs that target a large network of genes in T cells to restrict their cell cycle, memory formation and survival. Upon T cell activation, miR-15/16 are downregulated, allowing rapid expansion of differentiated effector T cells to mediate a sustained immune response. Here, using conditional deletion of miR-15/16 in immunosuppressive regulatory T cells (Tregs) that express FOXP3, we identify new functions of the miR-15/16 family in T cell immunity. miR-15/16 are indispensable to maintain peripheral tolerance by securing efficient suppression by a limited number of Tregs. miR-15/16-deficiency alters Treg expression of critical functional proteins including FOXP3, IL2Rα/CD25, CTLA4, PD-1 and IL7Rα/CD127, and results in accumulation of functionally impaired FOXP3loCD25loCD127hi Tregs. Excessive proliferation in the absence of miR-15/16 inhibition of cell cycle programs shifts Treg diversity and produces an effector Treg phenotype characterized by low expression of TCF1, CD25 and CD62L, and high expression of CD44. These Tregs fail to control immune activation of CD4+ effector T cells, leading to spontaneous multi-organ inflammation and increased allergic airway inflammation in a mouse model of asthma. Together, our results demonstrate that miR-15/16 expression in Tregs is essential to maintain immune tolerance.
RESUMEN
Demodex mites are commensal parasites of hair follicles (HFs). Normally asymptomatic, inflammatory outgrowth of mites can accompany malnutrition, immune dysfunction, and aging, but mechanisms restricting Demodex outgrowth are not defined. Here, we show that control of mite HF colonization in mice required group 2 innate lymphoid cells (ILC2s), interleukin-13 (IL-13), and its receptor, IL-4Ra-IL-13Ra1. HF-associated ILC2s elaborated IL-13 that attenuated HFs and epithelial proliferation at anagen onset; in their absence, Demodex colonization led to increased epithelial proliferation and replacement of gene programs for repair by aberrant inflammation, leading to the loss of barrier function and HF exhaustion. Humans with rhinophymatous acne rosacea, an inflammatory condition associated with Demodex, had increased HF inflammation with decreased type 2 cytokines, consistent with the inverse relationship seen in mice. Our studies uncover a key role for skin ILC2s and IL-13, which comprise an immune checkpoint that sustains cutaneous integrity and restricts pathologic infestation by colonizing HF mites.
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Infestaciones por Ácaros , Ácaros , Animales , Citocinas , Folículo Piloso/patología , Humanos , Inmunidad Innata , Inflamación , Interleucina-13 , Linfocitos/patología , Ratones , Infestaciones por Ácaros/complicaciones , Infestaciones por Ácaros/parasitología , Infestaciones por Ácaros/patología , SimbiosisAsunto(s)
Vértebras Cervicales , Prurito , Corticoesteroides , Gabapentina , Humanos , Prurito/tratamiento farmacológico , Prurito/etiologíaRESUMEN
Decades of work have elucidated cytokine signalling and transcriptional pathways that control T cell differentiation and have led the way to targeted biologic therapies that are effective in a range of autoimmune, allergic and inflammatory diseases. Recent evidence indicates that obesity and metabolic disease can also influence the immune system1-7, although the mechanisms and effects on immunotherapy outcomes remain largely unknown. Here, using two models of atopic dermatitis, we show that lean and obese mice mount markedly different immune responses. Obesity converted the classical type 2 T helper (TH2)-predominant disease associated with atopic dermatitis to a more severe disease with prominent TH17 inflammation. We also observed divergent responses to biologic therapies targeting TH2 cytokines, which robustly protected lean mice but exacerbated disease in obese mice. Single-cell RNA sequencing coupled with genome-wide binding analyses revealed decreased activity of nuclear receptor peroxisome proliferator-activated receptor-γ (PPARγ) in TH2 cells from obese mice relative to lean mice. Conditional ablation of PPARγ in T cells revealed that PPARγ is required to focus the in vivo TH response towards a TH2-predominant state and prevent aberrant non-TH2 inflammation. Treatment of obese mice with a small-molecule PPARγ agonist limited development of TH17 pathology and unlocked therapeutic responsiveness to targeted anti-TH2 biologic therapies. These studies reveal the effects of obesity on immunological disease and suggest a precision medicine approach to target the immune dysregulation caused by obesity.
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Dermatitis Atópica , PPAR gamma , Animales , Citocinas/metabolismo , Modelos Animales de Enfermedad , Inflamación/metabolismo , Ratones , Obesidad/metabolismo , PPAR gamma/agonistas , PPAR gamma/metabolismo , Medicina de Precisión , Análisis de Secuencia de ARN , Células Th2/metabolismoRESUMEN
BACKGROUND: mRNA SARS-CoV-2 vaccines are administered to 2 million individuals per day in the United States under US Food and Drug Administration emergency use authorization. METHODS: Observational cohort study of hospital employees who received their first SARS-CoV-2 mRNA vaccination between 14 December 2020 and 8 January 2021, including employees who reported onset of an injection site reaction ≥48 hours after administration of their first or second dose to an employee hotline. RESULTS: Thirteen female employees who received the mRNA-1273 vaccine (Moderna) during the first 3 weeks of the SARS-CoV-2 vaccine rollout at San Francisco General Hospital reported a pruritic rash at the injection site appearing 3 -9 days after receipt of their initial dose. Five had milder or similar reactions with earlier onset after the second dose. One additional female employee reported this delayed reaction only after the second dose. None reported serious adverse events or had symptoms severe enough to seek medical attention. These cases represented 1.1% of the 1275 female employees who received their first mRNA-1273 dose and 2.0% of the 557 who were aged 31 -45 years during this initial vaccine rollout. None of 675 males who initiated mRNA-1273 or 3612 employees of any sex who initiated BNT162b (Pfizer) vaccination during this period reported delayed-onset reactions. CONCLUSIONS: These results suggest that delayed-onset, injection site pruritic rashes after mRNA-1273 SARS-CoV-2 vaccine administration, lasting up to 1 week, occur commonly in females, do not lead to serious sequela, and should not deter receipt of the second vaccine dose.
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Vacuna nCoV-2019 mRNA-1273 , COVID-19 , Reacción en el Punto de Inyección/epidemiología , Vacuna nCoV-2019 mRNA-1273/efectos adversos , Adulto , COVID-19/epidemiología , COVID-19/prevención & control , Vacunas contra la COVID-19/efectos adversos , Estudios de Cohortes , Femenino , Hospitales , Humanos , Incidencia , Masculino , Persona de Mediana Edad , SARS-CoV-2 , Estados Unidos/epidemiologíaRESUMEN
Many skin manifestations of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection reflect activation of cutaneous and systemic immune responses involving effector pathways of both the innate and adaptive arms of the immune system. This article reviews evidence from the recent clinical and scientific literature that informs the current understanding of the consequences of coronavirus disease 2019 (COVID-19)-induced immune cell activation, as relevant to dermatology. Topics include the clinical consequences of autoantibody production in patients with COVID-19, immunologic evidence for chilblains as a manifestation of SARS-CoV-2 infection, and the relationship between type I interferons and COVID-19 disease severity.
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COVID-19/fisiopatología , Enfermedades de la Piel/fisiopatología , Eritema Pernio/fisiopatología , Eritema Multiforme/fisiopatología , Exantema/fisiopatología , Humanos , Pitiriasis Rosada/fisiopatología , Piel/fisiopatología , Enfermedades Cutáneas Vesiculoampollosas/fisiopatologíaRESUMEN
Pruritus is a common symptom of inflammatory skin conditions, including atopic dermatitis (AD). Although primary sensory neurons that transmit pruritic signals are well-cataloged, little is known about the neuronal alterations that occur as a result of skin disruption in AD. To address this question, we examined the molecular and behavioral consequences of challenging Grhl3PAR2/+ mice, which overexpress PAR2 in suprabasal keratinocytes, with serial topical application of the environmental allergen house dust mite (HDM). We monitored behavior and used RNA sequencing, qPCR, and in situ hybridization to evaluate gene expression in trigeminal ganglia (TG), before and after HDM. We found that neither Grhl3PAR2/+ nor wild-type (WT) mice exhibited spontaneous scratching, and pruritogen-induced acute scratching did not differ. In contrast, HDM exacerbated scratching in Grhl3PAR2/+ mice. Despite the absence of scratching in untreated Grhl3PAR2/+ mice, several TG genes in these mice were up-regulated compared to WT. HDM treatment of the Grhl3PAR2/+ mice enhanced up-regulation of this set of genes and induced additional genes, many within the subset of TG neurons that express TRPV1. The same set of genes was up-regulated in HDM-treated Grhl3PAR2/+ mice that did not scratch, but at lesser magnitude. Finally, we recorded comparable transcriptional changes in IL31Tg mice, demonstrating that a common genetic program is induced in two AD models. Taken together, we conclude that transcriptional changes that occur in primary sensory neurons in dermatitis-susceptible animals underlie a genetic priming that not only sensitizes the animal to chronic allergens but also contributes to pruritus in atopic skin disease.
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Alérgenos/toxicidad , Proteínas de Unión al ADN/fisiología , Dermatitis Atópica/patología , Receptor PAR-2/metabolismo , Células Receptoras Sensoriales/patología , Piel/patología , Factores de Transcripción/fisiología , Animales , Dermatitis Atópica/inducido químicamente , Dermatitis Atópica/metabolismo , Modelos Animales de Enfermedad , Ratones , Ratones Transgénicos , RNA-Seq , Receptor PAR-2/genética , Células Receptoras Sensoriales/efectos de los fármacos , Células Receptoras Sensoriales/metabolismo , Piel/efectos de los fármacos , Piel/inervación , Piel/metabolismoAsunto(s)
COVID-19/diagnóstico , SARS-CoV-2/aislamiento & purificación , Enfermedades de la Piel/diagnóstico , Enfermedades de la Piel/virología , Piel/virología , Adulto , COVID-19/inmunología , COVID-19/patología , COVID-19/virología , Femenino , Humanos , Masculino , Persona de Mediana Edad , Enfermedades de la Piel/inmunología , Enfermedades de la Piel/patología , Adulto JovenRESUMEN
Inflammatory response heterogeneity has impeded high-resolution dissection of diverse immune cell populations during activation. We characterize mouse cutaneous immune cells by single-cell RNA sequencing, after inducing inflammation using imiquimod and oxazolone dermatitis models. We identify 13 CD45+ subpopulations, which broadly represent most functionally characterized immune cell types. Oxazolone pervasively upregulates Jak2/Stat3 expression across T cells and antigen-presenting cells (APCs). Oxazolone also induces Il4/Il13 expression in newly infiltrating basophils, and Il4ra and Ccl24, most prominently in APCs. In contrast, imiquimod broadly upregulates Il17/Il22 and Ccl4/Ccl5. A comparative analysis of single-cell inflammatory transcriptional responses reveals that APC response to oxazolone is tightly restricted by cell identity, whereas imiquimod enforces shared programs on multiple APC populations in parallel. These global molecular patterns not only contrast immune responses on a systems level but also suggest that the mechanisms of new sources of inflammation can eventually be deduced by comparison to known signatures.
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Antiinflamatorios/administración & dosificación , Antineoplásicos Inmunológicos/efectos adversos , Erupciones por Medicamentos/etiología , Queratoacantoma/etiología , Anciano , Antineoplásicos Inmunológicos/administración & dosificación , Antígeno B7-H1/antagonistas & inhibidores , Erupciones por Medicamentos/tratamiento farmacológico , Femenino , Humanos , Inmunoterapia/efectos adversos , Inmunoterapia/métodos , Queratoacantoma/tratamiento farmacológico , Masculino , Receptor de Muerte Celular Programada 1/antagonistas & inhibidores , Resultado del TratamientoRESUMEN
Coordinate control of T cell proliferation, survival, and differentiation are essential for host protection from pathogens and cancer. Long-lived memory cells, whose precursors are formed during the initial immunological insult, provide protection from future encounters, and their generation is the goal of many vaccination strategies. microRNAs (miRNAs) are key nodes in regulatory networks that shape effective T cell responses through the fine-tuning of thousands of genes. Here, using compound conditional mutant mice to eliminate miR-15/16 family miRNAs in T cells, we show that miR-15/16 restrict T cell cycle, survival, and memory T cell differentiation. High throughput sequencing of RNA isolated by cross-linking immunoprecipitation of AGO2 combined with gene expression analysis in miR-15/16-deficient T cells indicates that these effects are mediated through the direct inhibition of an extensive network of target genes within pathways critical to cell cycle, survival, and memory.
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Ciclo Celular , Diferenciación Celular , Memoria Inmunológica , MicroARNs/metabolismo , Linfocitos T/citología , Linfocitos T/inmunología , Animales , Antígenos/metabolismo , Ciclo Celular/genética , Diferenciación Celular/genética , Supervivencia Celular/genética , Regulación de la Expresión Génica , Redes Reguladoras de Genes , Sitios Genéticos , Virus de la Coriomeningitis Linfocítica/fisiología , Ratones Transgénicos , MicroARNs/genéticaRESUMEN
Perturbations in the transcriptional programs specifying epidermal differentiation cause diverse skin pathologies ranging from impaired barrier function to inflammatory skin disease. However, the global scope and organization of this complex cellular program remain undefined. Here we report single-cell RNA sequencing profiles of 92,889 human epidermal cells from 9 normal and 3 inflamed skin samples. Transcriptomics-derived keratinocyte subpopulations reflect classic epidermal strata but also sharply compartmentalize epithelial functions such as cell-cell communication, inflammation, and WNT pathway modulation. In keratinocytes, â¼12% of assessed transcript expression varies in coordinate patterns, revealing undescribed gene expression programs governing epidermal homeostasis. We also identify molecular fingerprints of inflammatory skin states, including S100 activation in the interfollicular epidermis of normal scalp, enrichment of a CD1C+CD301A+ myeloid dendritic cell population in psoriatic epidermis, and IL1ßhiCCL3hiCD14+ monocyte-derived macrophages enriched in foreskin. This compendium of RNA profiles provides a critical step toward elucidating epidermal diseases of development, differentiation, and inflammation.
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Epidermis/metabolismo , Epidermis/patología , Inflamación/genética , Inflamación/patología , Análisis de la Célula Individual , Transcripción Genética , Anfirregulina/farmacología , Biomarcadores/metabolismo , Agregación Celular/genética , Comunicación Celular , Diferenciación Celular , Proliferación Celular , Prepucio/citología , Folículo Piloso/metabolismo , Humanos , Inflamación/inmunología , Queratinocitos/metabolismo , Cinética , Masculino , Psoriasis/genética , Psoriasis/inmunología , Psoriasis/patología , Proteínas S100/metabolismo , Factores de Tiempo , Transcriptoma/genética , Proteínas Wnt/metabolismoRESUMEN
Transgenic methods to manipulate CD4 T lymphocytes in vivo via forced expression of TCR transgenes and targeted "knockout" of individual genes by Cre-lox technology are fundamental to modern immunology. However, efforts to scale up functional analysis by modifying expression of larger numbers of genes in T cells ex vivo have proven surprisingly difficult. Early RNA interference experiments achieved successful small RNA transfection by using very high concentrations of short-interfering RNA (siRNA) [1], but primary T cells are generally resistant to standard electroporation, cationic liposome-, and calcium phosphate-mediated transfection methods. Moreover, although viral vectors can successfully introduce DNA fragments of varying length, expression of these constructs in primary T cells is low efficiency and the subcloning process laborious. In this context, the relatively recent discovery of dozens of highly expressed microRNAs (miRNAs) in the immune system provides both an opportunity and a new challenge [2, 3]. How can we query the miRNAome of a cell to assign particular roles to individual miRNAs? Here, we describe an optimized technique for efficient and reproducible transfection of primary mouse CD4 T cells in vitro with synthetic miRNA mimics.
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Hipersensibilidad/genética , MicroARNs/genética , Animales , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD4-Positivos/metabolismo , Electroporación , Expresión Génica , Hipersensibilidad/inmunología , Activación de Linfocitos , Ratones , Linfocitos T/inmunología , Linfocitos T/metabolismoRESUMEN
The transcription factor Foxp3 participates dominantly in the specification and function of Foxp3(+)CD4(+) regulatory T cells (T(reg) cells) but is neither strictly necessary nor sufficient to determine the characteristic T(reg) cell signature. Here we used computational network inference and experimental testing to assess the contribution of other transcription factors to this. Enforced expression of Helios or Xbp1 elicited distinct signatures, but Eos, IRF4, Satb1, Lef1 and GATA-1 elicited exactly the same outcome, acting in synergy with Foxp3 to activate expression of most of the T(reg) cell signature, including key transcription factors, and enhancing occupancy by Foxp3 at its genomic targets. Conversely, the T(reg) cell signature was robust after inactivation of any single cofactor. A redundant genetic switch thus 'locked in' the T(reg) cell phenotype, a model that would account for several aspects of T(reg) cell physiology, differentiation and stability.
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Linfocitos T Reguladores/inmunología , Linfocitos T Reguladores/metabolismo , Factores de Transcripción/metabolismo , Transcripción Genética , Animales , Antígenos CD4/biosíntesis , Diferenciación Celular , Biología Computacional , Proteínas de Unión al ADN/biosíntesis , Proteínas de Unión al ADN/genética , Factores de Transcripción Forkhead/genética , Factor de Transcripción GATA1/genética , Humanos , Factor de Transcripción Ikaros/biosíntesis , Factor de Transcripción Ikaros/genética , Factores Reguladores del Interferón/genética , Activación de Linfocitos , Factor de Unión 1 al Potenciador Linfoide/genética , Proteínas de Unión a la Región de Fijación a la Matriz/genética , Ratones , Ratones Endogámicos C57BL , Factores de Transcripción del Factor Regulador X , Serina Endopeptidasas/genética , Linfocitos T Reguladores/citología , Factores de Transcripción/biosíntesis , Factores de Transcripción/genética , Proteína 1 de Unión a la X-BoxRESUMEN
Immature thymocytes expressing autoreactive T-cell receptors (TCR) can adopt differing cell fates: clonal deletion by apoptosis or deviation into alternative lineages such as FoxP3(+) regulatory T cells (Treg). We revisited the role of the transcription factor Nr4a1 (Nur77), an immediate-early response gene induced by TCR engagement. Nr4a1KO mice show clear quantitative defects in antigen-induced clonal deletion. The impact of the Nr4a1 deletion is not enhanced by deletion of the proapoptotic factor Bim. In addition, Nr4a1 curtails initial differentiation into the Treg lineage in TCR transgenic mice and in nontransgenic mice. Transcriptional profiling of Nr4a1KO thymocytes under selection conditions reveals that Nr4a1 activates the transcription of several targets, consistent with these diverse actions: (i) Nr4a1 partakes in the induction of Bim after TCR triggering; (ii) perhaps paradoxically, Nr4a1 positively controls several transcripts of the Treg signature, in particular Ikzf2 and Tnfrsf9; (iii) consistent with its prosurvival and metabolic role in the liver, Nr4a1 is also required for the induction by TCR of a coordinated set of enzymes of the glycolytic and Krebs cycle pathways, which we propose may antagonize Treg selection as does activation of mTOR/Akt. Thus, Nr4a1 appears to act as a balancing molecule in fate determination at a critical juncture of T-cell differentiation.