RESUMEN
The mechanisms by which the number of memory CD8 T cells is stably maintained remains incompletely understood. It has been postulated that maintaining them requires help from CD4 T cells, because adoptively transferred memory CD8 T cells persist poorly in MHC class II (MHCII)-deficient mice. Here we show that chronic interferon-γ signals, not CD4 T cell-deficiency, are responsible for their attrition in MHCII-deficient environments. Excess IFN-γ is produced primarily by endogenous colonic CD8 T cells in MHCII-deficient mice. IFN-γ neutralization restores the number of memory CD8 T cells in MHCII-deficient mice, whereas repeated IFN-γ administration or transduction of a gain-of-function STAT1 mutant reduces their number in wild-type mice. CD127high memory cells proliferate actively in response to IFN-γ signals, but are more susceptible to attrition than CD127low terminally differentiated effector memory cells. Furthermore, single-cell RNA-sequencing of memory CD8 T cells reveals proliferating cells that resemble short-lived, terminal effector cells and documents global downregulation of gene signatures of long-lived memory cells in MHCII-deficient environments. We propose that chronic IFN-γ signals deplete memory CD8 T cells by compromising their long-term survival and by diverting self-renewing CD127high cells toward terminal differentiation.
Asunto(s)
Linfocitos T CD4-Positivos , Linfocitos T CD8-positivos , Memoria Inmunológica , Interferón gamma , Factor de Transcripción STAT1 , Animales , Linfocitos T CD8-positivos/inmunología , Interferón gamma/metabolismo , Interferón gamma/inmunología , Linfocitos T CD4-Positivos/inmunología , Ratones , Factor de Transcripción STAT1/metabolismo , Factor de Transcripción STAT1/genética , Factor de Transcripción STAT1/deficiencia , Ratones Endogámicos C57BL , Antígenos de Histocompatibilidad Clase II/inmunología , Antígenos de Histocompatibilidad Clase II/genética , Antígenos de Histocompatibilidad Clase II/metabolismo , Transducción de Señal , Ratones Noqueados , Células T de Memoria/inmunología , Células T de Memoria/metabolismo , Subunidad alfa del Receptor de Interleucina-7/metabolismo , Proliferación Celular , Traslado AdoptivoRESUMEN
Recent evidence indicates that viral components of the microbiota can contribute to intestinal homeostasis and protection from local inflammatory or infectious insults. However, host-derived mechanisms that regulate the virome remain largely unknown. In this study, we used colonization with the model commensal murine norovirus (MNV; strain CR6) to interrogate host-directed mechanisms of viral regulation, and we show that STAT1 is a central coordinator of both viral replication and antiviral T cell responses. In addition to restricting CR6 replication to the intestinal tract, we show that STAT1 regulates antiviral CD4+ and CD8+ T cell responses and prevents systemic viral-induced tissue damage and disease. Despite altered T cell responses that resemble those that mediate lethal immunopathology in systemic viral infections in STAT1-deficient mice, depletion of adaptive immune cells and their associated effector functions had no effect on CR6-induced disease. However, therapeutic administration of an antiviral compound limited viral replication, preventing virus-induced tissue damage and death without impacting the generation of inflammatory antiviral T cell responses. Collectively, our data show that STAT1 restricts MNV CR6 replication within the intestinal mucosa and that uncontrolled viral replication mediates disease rather than the concomitant development of dysregulated antiviral T cell responses in STAT1-deficient mice. IMPORTANCE The intestinal microbiota is a collection of bacteria, archaea, fungi, and viruses that colonize the mammalian gut. Coevolution of the host and microbiota has required development of immunological tolerance to prevent ongoing inflammatory responses against intestinal microbes. Breakdown of tolerance to bacterial components of the microbiota can contribute to immune activation and inflammatory disease. However, the mechanisms that are necessary to maintain tolerance to viral components of the microbiome, and the consequences of loss of tolerance, are less well understood. Here, we show that STAT1 is integral for preventing escape of a commensal-like virus, murine norovirus CR6 (MNV CR6), from the gut and that in the absence of STAT1, mice succumb to infection-induced disease. In contrast to the case with other systemic viral infections, mortality of STAT1-deficient mice is not driven by immune-mediated pathology. Our data demonstrate the importance of host-mediated geographical restriction of commensal-like viruses.
Asunto(s)
Infecciones por Caliciviridae , Norovirus , Factor de Transcripción STAT1 , Linfocitos T , Replicación Viral , Animales , Infecciones por Caliciviridae/mortalidad , Infecciones por Caliciviridae/fisiopatología , Mucosa Intestinal/virología , Ratones , Norovirus/fisiología , Factor de Transcripción STAT1/deficiencia , Factor de Transcripción STAT1/genética , Linfocitos T/inmunología , Linfocitos T/virologíaRESUMEN
BACKGROUND: Patients with Mendelian susceptibility to mycobacterial disease (MSMD) experience recurrent and/or persistent infectious diseases associated with poorly virulent mycobacteria. Multifocal osteomyelitis is among the representative manifestations of MSMD. The frequency of multifocal osteomyelitis is especially high in patients with MSMD etiologies that impair cellular response to IFN-γ, such as IFN-γR1, IFN-γR2, or STAT1 deficiency. OBJECTIVES: This study sought to characterize the mechanism underlying multifocal osteomyelitis in MSMD. METHODS: GM colonies prepared from bone marrow mononuclear cells from patients with autosomal dominant (AD) IFN-γR1 deficiency, AD STAT1 deficiency, or STAT1 gain of function (GOF) and from healthy controls were differentiated into osteoclasts in the presence or absence of IFN-γ. The inhibitory effect of IFN-γ on osteoclastogenesis was investigated by quantitative PCR, immunoblotting, tartrate-resistant acid phosphatase staining, and pit formation assays. RESULTS: Increased osteoclast numbers were identified by examining the histopathology of osteomyelitis in patients with AD IFN-γR1 deficiency or AD STAT1 deficiency. In the presence of receptor activator of nuclear factor kappa-B ligand and M-CSF, GM colonies from patients with AD IFN-γR1 deficiency, AD STAT1 deficiency, or STAT1 GOF differentiated into osteoclasts, similar to GM colonies from healthy volunteers. IFN-γ concentration-dependent inhibition of osteoclast formation was impaired in GM colonies from patients with AD IFN-γR1 deficiency or AD STAT1 deficiency, whereas it was enhanced in GM colonies from patients with STAT1 GOF. CONCLUSIONS: Osteoclast differentiation is increased in AD IFN-γR1 deficiency and AD STAT1 deficiency due to an impaired response to IFN-γ, leading to excessive osteoclast proliferation and, by inference, increased bone resorption in infected foci, which may underlie multifocal osteomyelitis.
Asunto(s)
Infecciones por Mycobacterium , Osteogénesis/efectos de los fármacos , Osteomielitis , Receptores de Interferón/deficiencia , Factor de Transcripción STAT1/deficiencia , Diferenciación Celular/efectos de los fármacos , Células Cultivadas , Predisposición Genética a la Enfermedad , Humanos , Interferón gamma/farmacología , Mutación , Infecciones por Mycobacterium/genética , Mycobacterium avium , Osteoclastos/efectos de los fármacos , Osteomielitis/genética , Receptores de Interferón/genética , Factor de Transcripción STAT1/genéticaRESUMEN
Dengue is one of the most prevalent arthropod-borne viral diseases in humans. There is still no effective vaccine or treatment to date. Previous studies showed that mosquito-derived factors present in saliva or salivary gland extract (SGE) contribute to the pathogenesis of dengue. In this study, we aimed to investigate the interplay between mosquito vector and DENV and to address the question of whether the mosquito vector alters the virus that leads to consequential disease manifestations in the mammalian host. DENV2 cultured in C6/36 cell line (culture-DENV2) was injected to Aedes aegypti intrathoracically. Saliva was collected from infected mosquitoes 7 days later. Exploiting the sensitivity of Stat1-/- mice to low dose of DENV2 delivered intradermally, we showed that DENV2 collected in infected mosquito saliva (msq-DENV2) induced more severe hemorrhage in mice than their culture counterpart. Msq-DENV2 was characterized by smaller particle size, larger plaque size and more rapid growth in mosquito as well as mammalian cell lines compared to culture-DENV2. In addition, msq-DENV2 was more efficient than culture-DENV2 in inducing Tnf mRNA production by mouse macrophage. Together, our results point to the possibility that the mosquito vector provides an environment that alters DENV2 by changing its growth characteristics as well as its potential to cause disease.
Asunto(s)
Aedes/virología , Virus del Dengue/fisiología , Mosquitos Vectores/virología , Factor de Transcripción STAT1/genética , Dengue Grave/genética , Aedes/fisiología , Animales , Línea Celular , Virus del Dengue/genética , Virus del Dengue/patogenicidad , Femenino , Humanos , Masculino , Ratones , Ratones Noqueados , Mosquitos Vectores/fisiología , Factor de Transcripción STAT1/deficiencia , Dengue Grave/metabolismo , Dengue Grave/virología , Virulencia , Replicación ViralRESUMEN
Signal transducer and activator of transcription 1 (STAT1) acts as a tumor suppressor molecule in colitis-associated colorectal cancer (CAC), particularly during the very early stages, modulating immune responses and controlling mechanisms such as apoptosis and cell proliferation. Previously, using an experimental model of CAC, we reported increased intestinal cell proliferation and faster tumor development, which were consistent with more signs of disease and damage, and reduced survival in STAT1-/- mice, compared with WT counterparts. However, the mechanisms through which STAT1 might prevent colorectal cancer progression preceded by chronic inflammation are still unclear. Here, we demonstrate that increased tumorigenicity related to STAT1 deficiency could be suppressed by IL-17 neutralization. The blockade of IL-17 in STAT1-/- mice reduced the accumulation of CD11b+Ly6ClowLy6G+ cells resembling granulocytic myeloid-derived suppressor cells (MDSCs) in both spleen and circulation. Additionally, IL-17 blockade reduced the recruitment of neutrophils into intestinal tissue, the expression and production of inflammatory cytokines, and the expression of intestinal STAT3. In addition, the anti-IL-17 treatment also reduced the expression of Arginase-1 and inducible nitric oxide synthase (iNOS) in the colon, both associated with the main suppressive activity of MDSCs. Thus, a lack of STAT1 signaling induces a significant change in the colonic microenvironment that supports inflammation and tumor formation. Anti-IL-17 treatment throughout the initial stages of CAC related to STAT1 deficiency abrogates the tumor formation possibly caused by myeloid cells.
Asunto(s)
Neoplasias Asociadas a Colitis/etiología , Granulocitos/patología , Interleucina-17/fisiología , Factor de Transcripción STAT1/fisiología , Animales , Anticuerpos Neutralizantes/administración & dosificación , Neoplasias Asociadas a Colitis/patología , Neoplasias Asociadas a Colitis/fisiopatología , Progresión de la Enfermedad , Femenino , Granulocitos/inmunología , Interleucina-17/antagonistas & inhibidores , Interleucina-17/inmunología , Ratones , Ratones Endogámicos BALB C , Ratones Noqueados , Células Supresoras de Origen Mieloide/inmunología , Células Supresoras de Origen Mieloide/patología , Neoplasias Experimentales/etiología , Neoplasias Experimentales/patología , Neoplasias Experimentales/fisiopatología , Factor de Transcripción STAT1/deficiencia , Factor de Transcripción STAT1/genética , Microambiente Tumoral/inmunologíaRESUMEN
Type I IFNs are so-named because they interfere with viral infection in vertebrate cells. The study of cellular responses to type I IFNs led to the discovery of the JAK-STAT signaling pathway, which also governs the response to other cytokine families. We review here the outcome of viral infections in mice and humans with engineered and inborn deficiencies, respectively, of (i) IFNAR1 or IFNAR2, selectively disrupting responses to type I IFNs, (ii) STAT1, STAT2, and IRF9, also impairing cellular responses to type II (for STAT1) and/or III (for STAT1, STAT2, IRF9) IFNs, and (iii) JAK1 and TYK2, also impairing cellular responses to cytokines other than IFNs. A picture is emerging of greater redundancy of human type I IFNs for protective immunity to viruses in natural conditions than was initially anticipated. Mouse type I IFNs are essential for protection against a broad range of viruses in experimental conditions. These findings suggest that various type I IFN-independent mechanisms of human cell-intrinsic immunity to viruses have yet to be discovered.
Asunto(s)
Predisposición Genética a la Enfermedad , Interferón Tipo I/metabolismo , Transducción de Señal , Virosis/etiología , Virosis/metabolismo , Alelos , Animales , Modelos Animales de Enfermedad , Genotipo , Humanos , Subunidad gamma del Factor 3 de Genes Estimulados por el Interferón/deficiencia , Interferones/metabolismo , Janus Quinasa 1/deficiencia , Síndrome de Job/genética , Ratones , Ratones Noqueados , Mutación , Fenotipo , Receptor de Interferón alfa y beta/genética , Receptor de Interferón alfa y beta/metabolismo , Factor de Transcripción STAT1/deficiencia , Factor de Transcripción STAT2/deficiencia , TYK2 Quinasa/deficiencia , TYK2 Quinasa/genéticaAsunto(s)
Genes Recesivos , Predisposición Genética a la Enfermedad , Trasplante de Células Madre Hematopoyéticas , Factor de Transcripción STAT1/deficiencia , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Manejo de la Enfermedad , Enfermedad Injerto contra Huésped/diagnóstico , Enfermedad Injerto contra Huésped/etiología , Trasplante de Células Madre Hematopoyéticas/efectos adversos , Trasplante de Células Madre Hematopoyéticas/métodos , Humanos , Acondicionamiento Pretrasplante/métodos , Resultado del TratamientoRESUMEN
Injured tubule epithelium stimulates a profibrotic milieu that accelerates loss of function in chronic kidney disease (CKD). This study tested the role of signal transducer and activator of transcription 1 (STAT1) in the progressive loss of kidney function in aristolochic acid (AA) nephropathy, a model of CKD. Mean serum creatinine concentration increased in wild-type (WT) littermates treated with AA, whereas Stat1-/- mice were protected. Focal increases in the apical expression of kidney injury molecule (KIM)-1 were observed in the proximal tubules of WT mice with AA treatment but were absent in Stat1-/- mice in the treatment group as well as in both control groups. A composite injury score, an indicator of proximal tubule injury, was reduced in Stat1-/- mice treated with AA. Increased expression of integrin-ß6 and phosphorylated Smad2/3 in proximal tubules as well as interstitial collagen and fibronectin were observed in WT mice following AA treatment but were all decreased in AA-treated Stat1-/- mice. The data indicated that STAT1 activation facilitated the development of progressive kidney injury and interstitial fibrosis in AA nephropathy.
Asunto(s)
Ácidos Aristolóquicos , Matriz Extracelular/metabolismo , Eliminación de Gen , Túbulos Renales Proximales/metabolismo , Insuficiencia Renal Crónica/prevención & control , Factor de Transcripción STAT1/deficiencia , Animales , Modelos Animales de Enfermedad , Matriz Extracelular/patología , Fibrosis , Receptor Celular 1 del Virus de la Hepatitis A/metabolismo , Cadenas beta de Integrinas/metabolismo , Túbulos Renales Proximales/patología , Masculino , Ratones Endogámicos C57BL , Ratones Noqueados , Fosforilación , Insuficiencia Renal Crónica/inducido químicamente , Insuficiencia Renal Crónica/metabolismo , Insuficiencia Renal Crónica/patología , Factor de Transcripción STAT1/genética , Transducción de Señal , Proteína Smad2/metabolismo , Proteína smad3/metabolismoAsunto(s)
Genes Dominantes , Predisposición Genética a la Enfermedad , Infecciones por Mycobacterium/diagnóstico , Infecciones por Mycobacterium/etiología , Linaje , Factor de Transcripción STAT1/deficiencia , Adulto , Alelos , Antituberculosos/uso terapéutico , Quimioterapia Combinada , Femenino , Estudios de Asociación Genética , Humanos , Lactante , Imagen por Resonancia Magnética , Masculino , Mutación , Infecciones por Mycobacterium/tratamiento farmacológico , Infecciones por Mycobacterium/prevención & control , Piel/patología , Resultado del Tratamiento , Tuberculosis Osteoarticular/diagnóstico , Tuberculosis Osteoarticular/etiologíaRESUMEN
Natural killer (NK) cells are important components of the innate immune defense against infections and cancers. Signal transducer and activator of transcription 1 (STAT1) is a transcription factor that is essential for NK cell maturation and NK cell-dependent tumor surveillance. Two alternatively spliced isoforms of STAT1 exist: a full-length STAT1α and a C-terminally truncated STAT1ß isoform. Aberrant splicing is frequently observed in cancer cells and several anti-cancer drugs interfere with the cellular splicing machinery. To investigate whether NK cell-mediated tumor surveillance is affected by a switch in STAT1 splicing, we made use of knock-in mice expressing either only the STAT1α (Stat1α/α) or the STAT1ß (Stat1ß/ß ) isoform. NK cells from Stat1α/α mice matured normally and controlled transplanted tumor cells as efficiently as NK cells from wild-type mice. In contrast, NK cells from Stat1ß/ß mice showed impaired maturation and effector functions, albeit less severe than NK cells from mice that completely lack STAT1 (Stat1-/- ). Mechanistically, we show that NK cell maturation requires the presence of STAT1α in the niche rather than in NK cells themselves and that NK cell maturation depends on IFNγ signaling under homeostatic conditions. The impaired NK cell maturation in Stat1ß/ß mice was paralleled by decreased IL-15 receptor alpha (IL-15Rα) surface levels on dendritic cells, macrophages and monocytes. Treatment of Stat1ß/ß mice with exogenous IL-15/IL-15Rα complexes rescued NK cell maturation but not their effector functions. Collectively, our findings provide evidence that STAT1 isoforms are not functionally redundant in regulating NK cell activity and that the absence of STAT1α severely impairs, but does not abolish, NK cell-dependent tumor surveillance.
Asunto(s)
Células Asesinas Naturales/citología , Linfopoyesis/fisiología , Factor de Transcripción STAT1/inmunología , Animales , Trasplante de Médula Ósea , Línea Celular Tumoral , Citotoxicidad Inmunológica , Vigilancia Inmunológica/efectos de los fármacos , Vigilancia Inmunológica/inmunología , Factor 3 de Genes Estimulados por el Interferón/deficiencia , Factor 3 de Genes Estimulados por el Interferón/genética , Factor 3 de Genes Estimulados por el Interferón/inmunología , Interleucina-15/farmacología , Subunidad alfa del Receptor de Interleucina-15 , Células Asesinas Naturales/efectos de los fármacos , Células Asesinas Naturales/inmunología , Depleción Linfocítica , Tejido Linfoide/citología , Linfoma/inmunología , Linfoma/patología , Linfopoyesis/efectos de los fármacos , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Especificidad de Órganos , Isoformas de Proteínas/genética , Isoformas de Proteínas/inmunología , Receptores de Interferón/deficiencia , Factor de Transcripción STAT1/deficiencia , Factor de Transcripción STAT1/genética , Organismos Libres de Patógenos Específicos , Bazo/citología , Receptor de Interferón gammaRESUMEN
Signal transducer and activator of transcription 1 (STAT1) is known to be an important player in inflammatory responses. STAT1 as a transcription factor regulates the expression of multiple proinflammatory genes. Inflammatory response is one of the common effects of ototoxicity. Our group reported that hair cells of STAT1 knockout (STAT1-KO) mice are less sensitive to ototoxic agents in-vitro. The effect of inflammatory responses in STAT1-KO mice has primarily been studied challenging them with several pathogens and analyzing different organs of those mice. However, the effect of STAT1 ablation in the mouse inner ear has not been reported. Therefore, we evaluated the cochlear function of wild type and STAT1-KO mice via auditory brain stem response (ABR) and performed histopathologic analysis of their temporal bones. We found ABR responses were affected in STAT1-KO mice with cases of bilateral and unilateral hearing impairment. Histopathologic examination of the middle and inner ears showed bilateral and unilateral otitis media. Otitis media was characterized by effusion of middle and inner ear that varied between the mice in volume and inflammatory cell content. In addition, the thickness of the middle ear mucosae in STAT1-KO mice were more pronounced than those in wild type mice. The degree of middle and inner ear inflammation correlated with ABR threshold elevation in STAT1-KO mice. It appears that a number of mice with inflammation underwent spontaneous resolution. The ABR thresholds were variable and showed a tendency to increase in homozygous and heterozygous STAT1-KO mice. These findings suggest that STAT1 ablation confers an increased susceptibility to otitis media leading to hearing impairment. Thus, the study supports the new role of STAT1 as otitis media predisposition gene.
Asunto(s)
Otitis Media/genética , Factor de Transcripción STAT1/genética , Animales , Cóclea/patología , Cóclea/fisiopatología , Oído Medio/patología , Oído Medio/fisiopatología , Potenciales Evocados Auditivos del Tronco Encefálico , Ratones , Ratones Endogámicos C57BL , Factor de Transcripción STAT1/deficienciaRESUMEN
An entirely plasmid-based reverse genetics (RG) system was recently developed for rotavirus (RV), opening new avenues for in-depth molecular dissection of RV biology, immunology, and pathogenesis. Several improvements to further optimize the RG efficiency have now been described. However, only a small number of individual RV strains have been recovered to date. None of the current methods have supported the recovery of murine RV, impeding the study of RV replication and pathogenesis in an in vivo suckling mouse model. Here, we describe useful modifications to the RG system that significantly improve rescue efficiency of multiple RV strains. In addition to the 11 group A RV segment-specific (+)RNAs [(+)ssRNAs], a chimeric plasmid was transfected, from which the capping enzyme NP868R of African swine fever virus (ASFV) and the T7 RNA polymerase were expressed. Second, a genetically modified MA104 cell line was used in which several components of the innate immunity were degraded. Using this RG system, we successfully recovered the simian RV RRV strain, the human RV CDC-9 strain, a reassortant between murine RV D6/2 and simian RV SA11 strains, and several reassortants and reporter RVs. All these recombinant RVs were rescued at a high efficiency (≥80% success rate) and could not be reliably rescued using several recently published RG strategies (<20%). This improved system represents an important tool and great potential for the rescue of other hard-to-recover RV strains such as low-replicating attenuated vaccine candidates or low-cell culture passage clinical isolates from humans or animals.IMPORTANCE Group A rotavirus (RV) remains as the single most important cause of severe acute gastroenteritis among infants and young children worldwide. An entirely plasmid-based reverse genetics (RG) system was recently developed, opening new ways for in-depth molecular study of RV. Despite several improvements to further optimize the RG efficiency, it has been reported that current strategies do not enable the rescue of all cultivatable RV strains. Here, we described a helpful modification to the current strategies and established a tractable RG system for the rescue of the simian RRV strain, the human CDC-9 strain, and a murine-like RV strain, which is suitable for both in vitro and in vivo studies. This improved RV reverse genetics system will facilitate study of RV biology in both in vitro and in vivo systems that will facilitate the improved design of RV vaccines, better antiviral therapies, and expression vectors.
Asunto(s)
Regulación Viral de la Expresión Génica , Interacciones Huésped-Patógeno/genética , Virus Reordenados/genética , Genética Inversa/métodos , Rotavirus/genética , Proteínas Virales/genética , Virus de la Fiebre Porcina Africana/genética , Virus de la Fiebre Porcina Africana/inmunología , Animales , Chlorocebus aethiops , ARN Polimerasas Dirigidas por ADN/genética , ARN Polimerasas Dirigidas por ADN/inmunología , Interacciones Huésped-Patógeno/inmunología , Humanos , Inmunidad Innata , Factores Reguladores del Interferón/deficiencia , Factores Reguladores del Interferón/genética , Factores Reguladores del Interferón/inmunología , Ratones , Nucleotidiltransferasas/genética , Nucleotidiltransferasas/inmunología , Plásmidos/química , Plásmidos/metabolismo , Caperuzas de ARN , Virus Reordenados/inmunología , Proteínas Recombinantes/genética , Proteínas Recombinantes/inmunología , Rotavirus/inmunología , Factor de Transcripción STAT1/deficiencia , Factor de Transcripción STAT1/genética , Factor de Transcripción STAT1/inmunología , Transfección , Células Vero , Proteínas Virales/inmunología , Replicación ViralRESUMEN
Autosomal recessive (AR) complete signal transducer and activator of transcription 1 (STAT1) deficiency is an extremely rare primary immunodeficiency that causes life-threatening mycobacterial and viral infections. Only seven patients from five unrelated families with this disorder have been so far reported. All causal STAT1 mutations reported are exonic and homozygous. We studied a patient with susceptibility to mycobacteria and virus infections, resulting in identification of AR complete STAT1 deficiency due to compound heterozygous mutations, both located in introns: c.128+2 T>G and c.542-8 A>G. Both mutations were the first intronic STAT1 mutations to cause AR complete STAT1 deficiency. Targeted RNA-seq documented the impairment of STAT1 mRNA expression and contributed to the identification of the intronic mutations. The patient's cells showed a lack of STAT1 expression and phosphorylation, and severe impairment of the cellular response to IFN-γ and IFN-α. The case reflects the importance of accurate clinical diagnosis and precise evaluation, to include intronic mutations, in the comprehensive genomic study when the patient lacks molecular pathogenesis. In conclusion, AR complete STAT1 deficiency can be caused by compound heterozygous and intronic mutations. Targeted RNA-seq-based systemic gene expression assay may help to increase diagnostic yield in inconclusive cases after comprehensive genomic study.
Asunto(s)
Enfermedades Genéticas Congénitas/genética , Factor de Transcripción STAT1/inmunología , Niño , Enfermedades Genéticas Congénitas/diagnóstico , Humanos , Masculino , Mutación , ARN Mensajero/genética , RNA-Seq , Factor de Transcripción STAT1/deficiencia , Factor de Transcripción STAT1/genéticaRESUMEN
Chikungunya virus (CHIKV), an emerging alphavirus, has infected millions of people. However, the factors modulating disease outcome remain poorly understood. Here, we show in germ-free mice or in oral antibiotic-treated conventionally housed mice with depleted intestinal microbiomes that greater CHIKV infection and spread occurs within 1 day of virus inoculation. Alteration of the microbiome alters TLR7-MyD88 signaling in plasmacytoid dendritic cells (pDCs) and blunts systemic production of type I interferon (IFN). Consequently, circulating monocytes express fewer IFN-stimulated genes and become permissive for CHIKV infection. Reconstitution with a single bacterial species, Clostridium scindens, or its derived metabolite, the secondary bile acid deoxycholic acid, can restore pDC- and MyD88-dependent type I IFN responses to restrict systemic CHIKV infection and transmission back to vector mosquitoes. Thus, symbiotic intestinal bacteria modulate antiviral immunity and levels of circulating alphaviruses within hours of infection through a bile acid-pDC-IFN signaling axis, which affects viremia, dissemination, and potentially transmission.
Asunto(s)
Ácidos y Sales Biliares/metabolismo , Fiebre Chikungunya/patología , Microbioma Gastrointestinal , Interferón Tipo I/metabolismo , Animales , Antibacterianos/farmacología , Fiebre Chikungunya/inmunología , Fiebre Chikungunya/veterinaria , Virus Chikungunya/genética , Virus Chikungunya/aislamiento & purificación , Clostridiales/fisiología , Células Dendríticas/citología , Células Dendríticas/inmunología , Células Dendríticas/metabolismo , Trasplante de Microbiota Fecal , Microbioma Gastrointestinal/efectos de los fármacos , Masculino , Glicoproteínas de Membrana/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Monocitos/citología , Monocitos/inmunología , Monocitos/metabolismo , Factor 88 de Diferenciación Mieloide/deficiencia , Factor 88 de Diferenciación Mieloide/genética , Factor 88 de Diferenciación Mieloide/metabolismo , ARN Viral/sangre , Factor de Transcripción STAT1/deficiencia , Transducción de Señal , Receptor Toll-Like 7/metabolismoRESUMEN
We demonstrate that female C57BL/6J mice are susceptible to a transient lower genital tract infection with MmuPV1 mouse papillomavirus and display focal histopathological abnormalities resembling those of human papillomavirus (HPV) infection. We took advantage of strains of genetically deficient mice to study in vivo the role of innate immune signaling in the control of papillomavirus. At 4 months, we sacrificed MmuPV1-infected mice and measured viral 757/3139 spliced transcripts by TaqMan reverse transcription-PCR (RT-PCR), localization of infection by RNAscope in situ hybridization, and histopathological abnormities by hematoxylin and eosin (H&E) staining. Among mice deficient in receptors for pathogen-associated molecular patterns, MyD88-/- and STING-/- mice had 1,350 and 80 copies of spliced transcripts/µg RNA, respectively, while no viral expression was detected in MAVS-/- and Ripk2-/- mice. Mice deficient in an adaptor molecule, STAT1-/-, for interferon signaling had 46,000 copies/µg RNA. Among mice with targeted deficiencies in the inflammatory response, interleukin-1 receptor knockout (IL-1R-/-) and caspase-1-/- mice had 350 and 30 copies/µg RNA, respectively. Among mice deficient in chemokine receptors, CCR6-/- mice had 120 copies/µg RNA, while CXCR2-/- and CXCR3-/- mice were negative. RNAscope confirmed focal infection in MyD88-/-, STAT1-/-, and CCR6-/- mice but was negative for other gene-deficient mice. Histological abnormalities were seen only in the latter mice. Our findings and the literature support a working model of innate immunity to papillomaviruses involving the activation of a MyD88-dependent pathway and IL-1 receptor signaling, control of viral replication by interferon-stimulated genes, and clearance of virus-transformed dysplastic cells by the action of the CCR6/CCL20 axis.IMPORTANCE Papillomaviruses infect stratified squamous epithelia, and the viral life cycle is linked to epithelial differentiation. Additionally, changes occur in viral and host gene expression, and immune cells are activated to modulate the infectious process. In vitro studies with keratinocytes cannot fully model the complex viral and host responses and do not reflect the contribution of local and migrating immune cells. We show that female C57BL/6J mice are susceptible to a transient papillomavirus cervicovaginal infection, and mice deficient in select genes involved in innate immune responses are susceptible to persistent infection with variable manifestations of histopathological abnormalities. The results of our studies support a working model of innate immunity to papillomaviruses, and the model provides a framework for more in-depth studies. A better understanding of mechanisms of early viral clearance and the development of approaches to induce clearance will be important for cancer prevention and the treatment of HPV-related diseases.
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Interacciones Huésped-Patógeno/inmunología , Factor 88 de Diferenciación Mieloide/inmunología , Papillomaviridae/inmunología , Infecciones por Papillomavirus/inmunología , ARN Mensajero/inmunología , ARN Viral/inmunología , Receptores Tipo I de Interleucina-1/inmunología , Proteínas Adaptadoras Transductoras de Señales/deficiencia , Proteínas Adaptadoras Transductoras de Señales/genética , Proteínas Adaptadoras Transductoras de Señales/inmunología , Empalme Alternativo , Animales , Caspasa 1/deficiencia , Caspasa 1/genética , Caspasa 1/inmunología , Cuello del Útero/inmunología , Cuello del Útero/virología , Femenino , Regulación de la Expresión Génica , Interacciones Huésped-Patógeno/genética , Humanos , Inmunidad Innata , Proteínas de la Membrana/deficiencia , Proteínas de la Membrana/genética , Proteínas de la Membrana/inmunología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Factor 88 de Diferenciación Mieloide/deficiencia , Factor 88 de Diferenciación Mieloide/genética , Papillomaviridae/crecimiento & desarrollo , Papillomaviridae/metabolismo , Infecciones por Papillomavirus/genética , Infecciones por Papillomavirus/virología , ARN Mensajero/genética , ARN Viral/genética , Proteína Serina-Treonina Quinasa 2 de Interacción con Receptor/deficiencia , Proteína Serina-Treonina Quinasa 2 de Interacción con Receptor/genética , Proteína Serina-Treonina Quinasa 2 de Interacción con Receptor/inmunología , Receptores CCR6/deficiencia , Receptores CCR6/genética , Receptores CCR6/inmunología , Receptores CXCR3/deficiencia , Receptores CXCR3/genética , Receptores CXCR3/inmunología , Receptores Tipo I de Interleucina-1/deficiencia , Receptores Tipo I de Interleucina-1/genética , Receptores de Interleucina-8B/deficiencia , Receptores de Interleucina-8B/genética , Receptores de Interleucina-8B/inmunología , Factor de Transcripción STAT1/deficiencia , Factor de Transcripción STAT1/genética , Factor de Transcripción STAT1/inmunología , Transducción de Señal , Vagina/inmunología , Vagina/virologíaRESUMEN
Interferon γ (IFN-γ) was shown to be a macrophage activating factor already in 1984. Consistently, inborn errors of IFN-γ immunity underlie Mendelian Susceptibility to Mycobacterial Disease (MSMD). MSMD is characterized by genetic predisposition to disease caused by weakly virulent mycobacterial species. Paradoxically, macrophages from patients with MSMD were little tested. Here, we report a disease modeling platform for studying IFN-γ related pathologies using macrophages derived from patient specific induced pluripotent stem cells (iPSCs). We used iPSCs from patients with autosomal recessive complete- and partial IFN-γR2 deficiency, partial IFN-γR1 deficiency and complete STAT1 deficiency. Macrophages from all patient iPSCs showed normal morphology and IFN-γ-independent functionality like phagocytic uptake of bioparticles and internalization of cytokines. For the IFN-γ-dependent functionalities, we observed that the deficiencies played out at various stages of the IFN-γ pathway, with the complete IFN-γR2 and complete STAT1 deficient cells showing the most severe phenotypes, in terms of upregulation of surface markers and induction of downstream targets. Although iPSC-derived macrophages with partial IFN-γR1 and IFN-γR2 deficiency still showed residual induction of downstream targets, they did not reduce the mycobacterial growth when challenged with Bacillus Calmette-Guérin. Taken together, we report a disease modeling platform to study the role of macrophages in patients with inborn errors of IFN-γ immunity.
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Células Madre Pluripotentes Inducidas/metabolismo , Interferón gamma/metabolismo , Macrófagos/metabolismo , Infecciones por Mycobacterium/genética , Mycobacterium , Receptores de Interferón/genética , Donantes de Sangre , Reprogramación Celular , Predisposición Genética a la Enfermedad , Humanos , Mutación , Infecciones por Mycobacterium/microbiología , Fenotipo , Receptores de Interferón/deficiencia , Factor de Transcripción STAT1/deficiencia , Factor de Transcripción STAT1/genética , Transducción de Señal/genética , Receptor de Interferón gammaRESUMEN
Cancers of the oral cavity remain the sixth most diagnosed cancer worldwide, with high rates of recurrence and mortality. We determined the role of STAT1 during oral carcinogenesis using two orthotopic models in mice genetically deficient for Stat1. Metastatic (LY2) and nonmetastatic (B4B8) head and neck squamous cell carcinoma (HNSCC) cell lines were injected into the oral cavity of Stat1 deficient (Stat1-/- ) and Stat1 competent (Stat1+/+ ) mice. Stat1-/- mice displayed increased tumor growth and metastasis compared to Stat1+/+ mice. Mechanistically, Stat1-/- mice displayed impaired CD4+ and CD8+ T-cell expansion compared to Stat1+/+ mice. This was associated with enhanced T-cell exhaustion, and severely attenuated T-cell antitumor effector responses including reduced expression of IFN-γ and perforin at the tumor site. Interestingly, tumor necrosis factor (TNF)-α production by T cells in tumor-bearing mice was suppressed by Stat1 deficiency. This deficiency in T-cell expansion and functional responses in mice was linked to PD-1 and CD69 overexpression in T cells of Stat1-/- mice. In contrast, we observed increased accumulation of CD11b+ Ly6G+ myeloid derived suppressor cells in tumors, draining lymph nodes, spleens and bone marrow of tumor-bearing Stat1-/- mice, resulting in a protumorigenic microenvironment. Our data demonstrates that STAT1 is an essential mediator of the antitumor response through inhibition of myeloid derived suppressor cell accumulation and promotion of T-cell mediated immune responses in murine head and neck squamous cell carcinoma. Selective induction of STAT1 phosphorylation in HNSCC patients could potentially improve oral tumor outcomes and response to therapy.
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Inmunomodulación , Células Supresoras de Origen Mieloide/inmunología , Células Supresoras de Origen Mieloide/metabolismo , Factor de Transcripción STAT1/metabolismo , Carcinoma de Células Escamosas de Cabeza y Cuello/etiología , Carcinoma de Células Escamosas de Cabeza y Cuello/metabolismo , Linfocitos T/inmunología , Linfocitos T/metabolismo , Animales , Línea Celular Tumoral , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Femenino , Humanos , Ganglios Linfáticos/patología , Masculino , Ratones , Ratones Noqueados , Metástasis de la Neoplasia , Estadificación de Neoplasias , Factor de Transcripción STAT1/deficiencia , Carcinoma de Células Escamosas de Cabeza y Cuello/patología , Subgrupos de Linfocitos T/inmunología , Subgrupos de Linfocitos T/metabolismo , Microambiente TumoralRESUMEN
Precision and personalized medicine is gaining importance in modern clinical medicine, as it aims to improve diagnostic precision and to reduce consequent therapeutic failures. In this regard, prior to use in human trials, animal models can help evaluate novel imaging approaches and therapeutic strategies and can help discover new biomarkers. Breast cancer is the most common malignancy in women worldwide, accounting for 25% of cases of all cancers and is responsible for approximately 500,000 deaths per year. Thus, it is important to identify accurate biomarkers for precise stratification of affected patients and for early detection of responsiveness to the selected therapeutic protocol. This review aims to summarize the latest advancements in preclinical molecular imaging in breast cancer mouse models. Positron emission tomography (PET) imaging remains one of the most common preclinical techniques used to evaluate biomarker expression in vivo, whereas magnetic resonance imaging (MRI), particularly diffusion-weighted (DW) sequences, has been demonstrated as capable of distinguishing responders from nonresponders for both conventional and innovative chemo- and immune-therapies with high sensitivity and in a noninvasive manner. The ability to customize therapies is desirable, as this will enable early detection of diseases and tailoring of treatments to individual patient profiles. Animal models remain irreplaceable in the effort to understand the molecular mechanisms and patterns of oncologic diseases.
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Neoplasias de la Mama/diagnóstico por imagen , Imagen Molecular/métodos , Medicina de Precisión/métodos , Radiofármacos , Ensayos Antitumor por Modelo de Xenoinjerto/métodos , Animales , Antineoplásicos/farmacocinética , Antineoplásicos/uso terapéutico , Neoplasias de la Mama/química , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/patología , Línea Celular Tumoral , Diagnóstico por Imagen/métodos , Resistencia a Antineoplásicos , Estrógenos , Femenino , Predicción , Humanos , Ratones , Ratones Noqueados , Proteínas de Neoplasias/análisis , Trasplante de Neoplasias , Neoplasias Hormono-Dependientes/diagnóstico por imagen , Neoplasias Hormono-Dependientes/patología , Progesterona , Receptor ErbB-2/análisis , Proteínas Recombinantes de Fusión , Factor de Transcripción STAT1/deficiencia , Sensibilidad y Especificidad , Trastuzumab/farmacocinética , Trastuzumab/uso terapéuticoRESUMEN
BACKGROUND & AIMS: Interferon lambda (IFNL) is expressed at high levels by intestinal epithelial cells (IECs) and mucosal immune cells in response to infection and inflammation. We investigated whether IFNL might contribute to pathogenesis of Crohn's disease (CD). METHODS: We obtained serum samples and terminal ileum biopsies from 47 patients with CD and 16 healthy individuals (controls). We measured levels of IFNL by enzyme-linked immunosorbent assay and immunohistochemistry and location of expression by confocal microscopy. Activation of IFNL signaling via STAT1 was measured in areas of no, mild, moderate, and severe inflammation and correlated with Paneth cell homeostasis and inflammation. IFNL expression and function were studied in wild-type mice and mice with intestinal epithelial cell-specific (ΔIEC) disruption or full-body disruption of specific genes (Mlkl-/-, Stat1ΔIEC, Casp8ΔIEC, Casp8ΔIECRipk3-/-, Casp8ΔIECTnfr-/-, Casp8ΔIECMlkl-/-, and Nod2-/- mice). Some mice were given tail vein injections of a vector encoding a secreted form of IFNL. Intestinal tissues were collected from mice and analyzed by immunohistochemistry and immunoblots. We generated 3-dimensional small intestinal organoids from mice and studied the effects of IFNL and inhibitors of STAT-signaling pathway. RESULTS: Patients with CD had significant increases in serum and ileal levels of IFNL compared with controls. Levels of IFNL were highest in ileum tissues with severe inflammation. High levels of IFNL associated with a reduced number of Paneth cells and increased cell death at the crypt bottom in inflamed ileum samples. Intestinal tissues from the ileum of wild-type mice injected with a vector expressing IFNL had reduced numbers of Paneth cells. IFNL-induced death of Paneth cells in mice did not occur via apoptosis, but required Mixed Lineage Kinase Domain Like (MLKL) and activation of Signal transducer and activator of transcription 1 (STAT1). In organoids, inhibitors of Janus kinase (JAK) signaling via STAT1 (glucocorticoids, tofacitinib, or filgotinib) reduced expression of proteins that mediate cell death and prevented Paneth cell death. CONCLUSIONS: Levels of IFNL are increased in serum and inflamed ileal tissues from patients with CD and associated with a loss of Paneth cells. Expression of a secreted form of IFNL in mice results in loss of Paneth cells from intestinal tissues, via STAT1 and MLKL, controlled by caspase 8. Strategies to reduce IFNL or block its effects might be developed for treatment of patients with CD affecting the terminal ileum.
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Enfermedad de Crohn/metabolismo , Íleon/metabolismo , Interferones/metabolismo , Interleucinas/metabolismo , Células de Paneth/metabolismo , Factor de Transcripción STAT1/metabolismo , Animales , Caspasa 8/genética , Caspasa 8/metabolismo , Muerte Celular , Enfermedad de Crohn/inmunología , Enfermedad de Crohn/patología , Modelos Animales de Enfermedad , Humanos , Íleon/inmunología , Íleon/patología , Interferones/genética , Interleucinas/genética , Ratones Endogámicos C57BL , Ratones Noqueados , Células de Paneth/inmunología , Células de Paneth/patología , Proteínas Quinasas/genética , Proteínas Quinasas/metabolismo , Factor de Transcripción STAT1/deficiencia , Factor de Transcripción STAT1/genética , Transducción de Señal , Técnicas de Cultivo de Tejidos , Regulación hacia ArribaRESUMEN
A signal transducer and activator of transcription-1-deficient patient presented with prolonged fever, cachexia, anemia, hypoalbuminemia and finally relapsing debilitating mycobacterial osteomyelitis while receiving a previously effective antimycobacterial treatment. Progression despite rigorous workup and multiple antibiotics prompted shotgun metagenomics revealing adenovirus in liver samples. Brincidofovir led to a complete, sustained clinical recovery, including osteomyelitis, probably attributed to reversal of adenovirus-induced immune dysregulation.