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1.
Nat Immunol ; 21(1): 86-100, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31844327

RESUMO

By developing a high-density murine immunophenotyping platform compatible with high-throughput genetic screening, we have established profound contributions of genetics and structure to immune variation (http://www.immunophenotype.org). Specifically, high-throughput phenotyping of 530 unique mouse gene knockouts identified 140 monogenic 'hits', of which most had no previous immunologic association. Furthermore, hits were collectively enriched in genes for which humans show poor tolerance to loss of function. The immunophenotyping platform also exposed dense correlation networks linking immune parameters with each other and with specific physiologic traits. Such linkages limit freedom of movement for individual immune parameters, thereby imposing genetically regulated 'immunologic structures', the integrity of which was associated with immunocompetence. Hence, we provide an expanded genetic resource and structural perspective for understanding and monitoring immune variation in health and disease.


Assuntos
Infecções por Enterobacteriaceae/imunologia , Variação Genética/genética , Ensaios de Triagem em Larga Escala/métodos , Imunofenotipagem/métodos , Infecções por Salmonella/imunologia , Animais , Citrobacter/imunologia , Infecções por Enterobacteriaceae/microbiologia , Feminino , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Modelos Animais , Salmonella/imunologia , Infecções por Salmonella/microbiologia
2.
Nat Immunol ; 17(9): 1046-56, 2016 09.
Artigo em Inglês | MEDLINE | ID: mdl-27478939

RESUMO

Single-nucleotide variations in C13orf31 (LACC1) that encode p.C284R and p.I254V in a protein of unknown function (called 'FAMIN' here) are associated with increased risk for systemic juvenile idiopathic arthritis, leprosy and Crohn's disease. Here we set out to identify the biological mechanism affected by these coding variations. FAMIN formed a complex with fatty acid synthase (FASN) on peroxisomes and promoted flux through de novo lipogenesis to concomitantly drive high levels of fatty-acid oxidation (FAO) and glycolysis and, consequently, ATP regeneration. FAMIN-dependent FAO controlled inflammasome activation, mitochondrial and NADPH-oxidase-dependent production of reactive oxygen species (ROS), and the bactericidal activity of macrophages. As p.I254V and p.C284R resulted in diminished function and loss of function, respectively, FAMIN determined resilience to endotoxin shock. Thus, we have identified a central regulator of the metabolic function and bioenergetic state of macrophages that is under evolutionary selection and determines the risk of inflammatory and infectious disease.


Assuntos
Artrite Juvenil/genética , Doença de Crohn/genética , Infecções/genética , Hanseníase/genética , Macrófagos/imunologia , Proteínas/genética , Choque Séptico/genética , Trifosfato de Adenosina/metabolismo , Animais , Bacteriólise , Células Cultivadas , Metabolismo Energético , Ácido Graxo Sintase Tipo I/metabolismo , Predisposição Genética para Doença , Humanos , Inflamassomos/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular , Metabolismo dos Lipídeos/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , NADPH Oxidases/metabolismo , Oxirredução , Polimorfismo de Nucleotídeo Único , Risco
3.
Immunity ; 50(4): 1099-1114.e10, 2019 04 16.
Artigo em Inglês | MEDLINE | ID: mdl-30876876

RESUMO

Inflammatory bowel disease is a chronic, relapsing condition with two subtypes, Crohn's disease (CD) and ulcerative colitis (UC). Genome-wide association studies (GWASs) in UC implicate a FCGR2A variant that alters the binding affinity of the antibody receptor it encodes, FcγRIIA, for immunoglobulin G (IgG). Here, we aimed to understand the mechanisms whereby changes in FcγRIIA affinity would affect inflammation in an IgA-dominated organ. We found a profound induction of anti-commensal IgG and a concomitant increase in activating FcγR signaling in the colonic mucosa of UC patients. Commensal-IgG immune complexes engaged gut-resident FcγR-expressing macrophages, inducing NLRP3- and reactive-oxygen-species-dependent production of interleukin-1ß (IL-1ß) and neutrophil-recruiting chemokines. These responses were modulated by the FCGR2A genotype. In vivo manipulation of macrophage FcγR signal strength in a mouse model of UC determined the magnitude of intestinal inflammation and IL-1ß-dependent type 17 immunity. The identification of an important contribution of IgG-FcγR-dependent inflammation to UC has therapeutic implications.


Assuntos
Anticorpos Antibacterianos/imunologia , Colite Ulcerativa/imunologia , Microbioma Gastrointestinal/imunologia , Imunoglobulina G/imunologia , Interleucina-1beta/imunologia , Células Th17/imunologia , Animais , Colite/induzido quimicamente , Colite/imunologia , Colite/microbiologia , Colite/patologia , Colite Ulcerativa/microbiologia , Colite Ulcerativa/patologia , Sulfato de Dextrana/toxicidade , Regulação da Expressão Gênica , Genótipo , Humanos , Inflamação , Interleucina-8/biossíntese , Interleucina-8/genética , Mucosa Intestinal/imunologia , Mucosa Intestinal/microbiologia , Macrófagos/imunologia , Camundongos , Fagócitos/imunologia , RNA Mensageiro/biossíntese , Espécies Reativas de Oxigênio , Receptores de IgG/biossíntese , Receptores de IgG/genética , Receptores de IgG/imunologia
4.
Cell ; 154(2): 452-64, 2013 Jul 18.
Artigo em Inglês | MEDLINE | ID: mdl-23870131

RESUMO

Mutations in whole organisms are powerful ways of interrogating gene function in a realistic context. We describe a program, the Sanger Institute Mouse Genetics Project, that provides a step toward the aim of knocking out all genes and screening each line for a broad range of traits. We found that hitherto unpublished genes were as likely to reveal phenotypes as known genes, suggesting that novel genes represent a rich resource for investigating the molecular basis of disease. We found many unexpected phenotypes detected only because we screened for them, emphasizing the value of screening all mutants for a wide range of traits. Haploinsufficiency and pleiotropy were both surprisingly common. Forty-two percent of genes were essential for viability, and these were less likely to have a paralog and more likely to contribute to a protein complex than other genes. Phenotypic data and more than 900 mutants are openly available for further analysis. PAPERCLIP:


Assuntos
Técnicas Genéticas , Camundongos Knockout , Fenótipo , Animais , Doença/genética , Modelos Animais de Doenças , Feminino , Genes Essenciais , Estudo de Associação Genômica Ampla , Masculino , Camundongos
5.
Nature ; 595(7865): 96-100, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-34040257

RESUMO

Trypanosomes are protozoan parasites that cause infectious diseases, including African trypanosomiasis (sleeping sickness) in humans and nagana in economically important livestock1,2. An effective vaccine against trypanosomes would be an important control tool, but the parasite has evolved sophisticated immunoprotective mechanisms-including antigenic variation3-that present an apparently insurmountable barrier to vaccination. Here we show, using a systematic genome-led vaccinology approach and a mouse model of Trypanosoma vivax infection4, that protective invariant subunit vaccine antigens can be identified. Vaccination with a single recombinant protein comprising the extracellular region of a conserved cell-surface protein that is localized to the flagellum membrane (which we term 'invariant flagellum antigen from T. vivax') induced long-lasting protection. Immunity was passively transferred with immune serum, and recombinant monoclonal antibodies to this protein could induce sterile protection and revealed several mechanisms of antibody-mediated immunity, including a major role for complement. Our discovery identifies a vaccine candidate for an important parasitic disease that has constrained socioeconomic development in countries in sub-Saharan Africa5, and provides evidence that highly protective vaccines against trypanosome infections can be achieved.


Assuntos
Antígenos de Protozoários/imunologia , Vacinas Protozoárias/imunologia , Trypanosoma vivax/imunologia , Tripanossomíase Africana/imunologia , Tripanossomíase Africana/prevenção & controle , Animais , Antígenos de Protozoários/química , Proteínas do Sistema Complemento/imunologia , Sequência Conservada/imunologia , Modelos Animais de Doenças , Feminino , Flagelos/química , Flagelos/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Vacinas Protozoárias/química , Fatores de Tempo , Trypanosoma vivax/química , Trypanosoma vivax/citologia , Tripanossomíase Africana/parasitologia , Vacinas de Subunidades Antigênicas/química , Vacinas de Subunidades Antigênicas/imunologia
6.
Nature ; 587(7834): 472-476, 2020 11.
Artigo em Inglês | MEDLINE | ID: mdl-33149302

RESUMO

The central nervous system has historically been viewed as an immune-privileged site, but recent data have shown that the meninges-the membranes that surround the brain and spinal cord-contain a diverse population of immune cells1. So far, studies have focused on macrophages and T cells, but have not included a detailed analysis of meningeal humoral immunity. Here we show that, during homeostasis, the mouse and human meninges contain IgA-secreting plasma cells. These cells are positioned adjacent to dural venous sinuses: regions of slow blood flow with fenestrations that can potentially permit blood-borne pathogens to access the brain2. Peri-sinus IgA plasma cells increased with age and following a breach of the intestinal barrier. Conversely, they were scarce in germ-free mice, but their presence was restored by gut re-colonization. B cell receptor sequencing confirmed that meningeal IgA+ cells originated in the intestine. Specific depletion of meningeal plasma cells or IgA deficiency resulted in reduced fungal entrapment in the peri-sinus region and increased spread into the brain following intravenous challenge, showing that meningeal IgA is essential for defending the central nervous system at this vulnerable venous barrier surface.


Assuntos
Cavidades Cranianas/imunologia , Microbioma Gastrointestinal/imunologia , Imunoglobulina A Secretora/imunologia , Intestinos/imunologia , Meninges/imunologia , Plasmócitos/imunologia , Idoso , Envelhecimento/imunologia , Animais , Barreira Hematoencefálica/imunologia , Feminino , Fungos/imunologia , Vida Livre de Germes , Humanos , Intestinos/citologia , Intestinos/microbiologia , Masculino , Meninges/irrigação sanguínea , Meninges/citologia , Camundongos , Camundongos Endogâmicos C57BL , Plasmócitos/citologia
7.
J Immunol ; 210(5): 547-557, 2023 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-36637239

RESUMO

Prolidase deficiency (PD) is a multisystem disorder caused by mutations in the PEPD gene, which encodes a ubiquitously expressed metallopeptidase essential for the hydrolysis of dipeptides containing C-terminal proline or hydroxyproline. PD typically presents in childhood with developmental delay, skin ulcers, recurrent infections, and, in some patients, autoimmune features that can mimic systemic lupus erythematosus. The basis for the autoimmune association is uncertain, but might be due to self-antigen exposure with tissue damage, or indirectly driven by chronic infection and microbial burden. In this study, we address the question of causation and show that Pepd-null mice have increased antinuclear autoantibodies and raised serum IgA, accompanied by kidney immune complex deposition, consistent with a systemic lupus erythematosus-like disease. These features are associated with an accumulation of CD4 and CD8 effector T cells in the spleen and liver. Pepd deficiency leads to spontaneous T cell activation and proliferation into the effector subset, which is cell intrinsic and independent of Ag receptor specificity or antigenic stimulation. However, an increase in KLRG1+ effector CD8 cells is not observed in mixed chimeras, in which the autoimmune phenotype is also absent. Our findings link autoimmune susceptibility in PD to spontaneous T cell dysfunction, likely to be acting in combination with immune activators that lie outside the hemopoietic system but result from the abnormal metabolism or loss of nonenzymatic prolidase function. This knowledge provides insight into the role of prolidase in the maintenance of self-tolerance and highlights the importance of treatment to control T cell activation.


Assuntos
Lúpus Eritematoso Sistêmico , Deficiência de Prolidase , Animais , Camundongos , Autoimunidade , Ativação Linfocitária , Autoantígenos
8.
PLoS Pathog ; 18(2): e1010364, 2022 02.
Artigo em Inglês | MEDLINE | ID: mdl-35202447

RESUMO

Leishmaniasis is an infectious disease caused by protozoan parasites belonging to the genus Leishmania for which there are no approved human vaccines. Infections localise to different tissues in a species-specific manner with the visceral form of the disease caused by Leishmania donovani and L. infantum being the most deadly in humans. Although Leishmania spp. parasites are predominantly intracellular, the visceral disease can be prevented in dogs by vaccinating with a complex mixture of secreted products from cultures of L. infantum promastigotes. With the logic that extracellular parasite proteins make good subunit vaccine candidates because they are directly accessible to vaccine-elicited host antibodies, here we attempt to discover proteins that are essential for in vitro growth and host infection with the goal of identifying subunit vaccine candidates. Using an in silico analysis of the Leishmania donovani genome, we identified 92 genes encoding proteins that are predicted to be secreted or externally anchored to the parasite membrane by a single transmembrane region or a GPI anchor. By selecting a transgenic L. donovani parasite that expresses both luciferase and the Cas9 nuclease, we systematically attempted to target all 92 genes by CRISPR genome editing and identified four that were required for in vitro growth. For fifty-five genes, we infected cohorts of mice with each mutant parasite and by longitudinally quantifying parasitaemia with bioluminescent imaging, showed that nine genes had evidence of an attenuated infection although all ultimately established an infection. Finally, we expressed two genes as full-length soluble recombinant proteins and tested them as subunit vaccine candidates in a murine preclinical infection model. Both proteins elicited significant levels of protection against the uncontrolled development of a splenic infection warranting further investigation as subunit vaccine candidates against this deadly infectious tropical disease.


Assuntos
Leishmania donovani , Leishmania infantum , Leishmaniose Visceral , Leishmaniose , Parasitos , Animais , Cães , Leishmania donovani/genética , Camundongos
9.
PLoS Pathog ; 17(1): e1009224, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-33481935

RESUMO

Animal African trypanosomiasis (AAT) is a severe, wasting disease of domestic livestock and diverse wildlife species. The disease in cattle kills millions of animals each year and inflicts a major economic cost on agriculture in sub-Saharan Africa. Cattle AAT is caused predominantly by the protozoan parasites Trypanosoma congolense and T. vivax, but laboratory research on the pathogenic stages of these organisms is severely inhibited by difficulties in making even minor genetic modifications. As a result, many of the important basic questions about the biology of these parasites cannot be addressed. Here we demonstrate that an in vitro culture of the T. congolense genomic reference strain can be modified directly in the bloodstream form reliably and at high efficiency. We describe a parental single marker line that expresses T. congolense-optimized T7 RNA polymerase and Tet repressor and show that minichromosome loci can be used as sites for stable, regulatable transgene expression with low background in non-induced cells. Using these tools, we describe organism-specific constructs for inducible RNA-interference (RNAi) and demonstrate knockdown of multiple essential and non-essential genes. We also show that a minichromosomal site can be exploited to create a stable bloodstream-form line that robustly provides >40,000 independent stable clones per transfection-enabling the production of high-complexity libraries of genome-scale. Finally, we show that modified forms of T. congolense are still infectious, create stable high-bioluminescence lines that can be used in models of AAT, and follow the course of infections in mice by in vivo imaging. These experiments establish a base set of tools to change T. congolense from a technically challenging organism to a routine model for functional genetics and allow us to begin to address some of the fundamental questions about the biology of this important parasite.


Assuntos
Genética Microbiana , Proteínas de Protozoários/genética , Transgenes , Trypanosoma congolense/genética , Trypanosoma congolense/patogenicidade , Tripanossomíase Africana/parasitologia , Animais , Feminino , Genoma de Protozoário , Técnicas In Vitro , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Tripanossomíase Africana/genética
10.
Nature ; 541(7636): 233-236, 2017 01 12.
Artigo em Inglês | MEDLINE | ID: mdl-28052056

RESUMO

Metastasis is the leading cause of death for cancer patients. This multi-stage process requires tumour cells to survive in the circulation, extravasate at distant sites, then proliferate; it involves contributions from both the tumour cell and tumour microenvironment ('host', which includes stromal cells and the immune system). Studies suggest the early steps of the metastatic process are relatively efficient, with the post-extravasation regulation of tumour growth ('colonization') being critical in determining metastatic outcome. Here we show the results of screening 810 mutant mouse lines using an in vivo assay to identify microenvironmental regulators of metastatic colonization. We identify 23 genes that, when disrupted in mouse, modify the ability of tumour cells to establish metastatic foci, with 19 of these genes not previously demonstrated to play a role in host control of metastasis. The largest reduction in pulmonary metastasis was observed in sphingosine-1-phosphate (S1P) transporter spinster homologue 2 (Spns2)-deficient mice. We demonstrate a novel outcome of S1P-mediated regulation of lymphocyte trafficking, whereby deletion of Spns2, either globally or in a lymphatic endothelial-specific manner, creates a circulating lymphopenia and a higher percentage of effector T cells and natural killer (NK) cells present in the lung. This allows for potent tumour cell killing, and an overall decreased metastatic burden.


Assuntos
Proteínas de Transporte de Ânions/genética , Proteínas de Transporte de Ânions/metabolismo , Genoma/genética , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/secundário , Metástase Neoplásica/genética , Metástase Neoplásica/patologia , Animais , Proteínas de Transporte de Ânions/deficiência , Linhagem Celular Tumoral , Movimento Celular , Modelos Animais de Doenças , Feminino , Genômica , Células Matadoras Naturais/citologia , Células Matadoras Naturais/imunologia , Neoplasias Pulmonares/imunologia , Neoplasias Pulmonares/patologia , Linfopenia/genética , Linfopenia/patologia , Lisofosfolipídeos/metabolismo , Masculino , Camundongos , Esfingosina/análogos & derivados , Esfingosina/metabolismo , Linfócitos T/citologia , Linfócitos T/imunologia , Microambiente Tumoral
11.
Br J Nurs ; 32(7): S4-S10, 2023 Apr 06.
Artigo em Inglês | MEDLINE | ID: mdl-37027407

RESUMO

Introduction: The widespread adoption of the ANTT®Clinical Practice Framework as a single standard for aseptic technique, has highlighted that many clinical procedures do not require a sterile procedure pack to be performed safely and aseptically. This study explores the utilisation of a partially-sterile procedure pack that is specifically tailored to Standard-ANTT. Methods: A prospective project improvement evaluation, using a non-paired sample (pre: n=41; post: n =33) of emergency department staff in an NHS hospital. Staff were evaluated performing peripheral intravenous cannulations (PIVC) using Standard-ANTT and the B. Braun Standard-ANTT peripheral cannulation pack. Findings: Significant improvements were observed in practice following the implementation of the pack and training in Standard-ANTT, including: Key-Part protection significantly improved (pre: n=28, 68.2%; post: n=33, 100%), and reduction in the Key-Site being touched after disinfection (pre: n=17; 41.4%; post n=5; 15.1%). Conclusions: In conjunction with appropriate education and training, this study provides proof of concept that due to the widespread use of the ANTT Clinical Practice Framework as a single standard aseptic technique, procedure packs that are specifically tailored to Standard-ANTT, can help to promote best practice and improve efficiencies. DEFINITIONS: Partially-sterile procedure pack - all items required to be sterile remain in their individual blister wrapper. The final assembled pack itself is not then subjected to a further round of sterilisation as it is not needed. Sterile procedure pack - often contains a mixture of non-sterile and sterile items that have been stripped from their individual blister wrapper requiring the sterilisation of the final assembled pack.


Assuntos
Vesícula , Cateterismo Periférico , Humanos , Estudos Prospectivos , Bandagens , Hospitais
12.
J Infect Dis ; 225(8): 1435-1446, 2022 04 19.
Artigo em Inglês | MEDLINE | ID: mdl-32524140

RESUMO

BACKGROUND: Schistosomiasis is a major global health problem caused by blood-dwelling parasitic worms, which is currently tackled primarily by mass administration of the drug praziquantel. Appropriate drug treatment strategies are informed by diagnostics that establish the prevalence and intensity of infection, which, in regions of low transmission, should be highly sensitive. METHODS: To identify sensitive new serological markers of Schistosoma mansoni infections, we have compiled a recombinant protein library of parasite cell-surface and secreted proteins expressed in mammalian cells. RESULTS: Together with a time series of sera samples from volunteers experimentally infected with a defined number of male parasites, we probed this protein library to identify several markers that can detect primary infections with as low as 10 parasites and as early as 5 weeks postinfection. CONCLUSIONS: These new markers could be further explored as valuable tools to detect ongoing and previous S mansoni infections, including in endemic regions where transmission is low.


Assuntos
Esquistossomose mansoni , Esquistossomose , Animais , Biomarcadores , Humanos , Masculino , Mamíferos , Camundongos , Praziquantel/uso terapêutico , Proteínas Recombinantes , Schistosoma mansoni , Esquistossomose/tratamento farmacológico , Esquistossomose mansoni/diagnóstico , Esquistossomose mansoni/tratamento farmacológico , Esquistossomose mansoni/parasitologia
13.
Nature ; 539(7627): 102-106, 2016 11 03.
Artigo em Inglês | MEDLINE | ID: mdl-27749818

RESUMO

Innate lymphoid cells (ILCs) functionally resemble T lymphocytes in cytotoxicity and cytokine production but lack antigen-specific receptors, and they are important regulators of immune responses and tissue homeostasis. ILCs are generated from common lymphoid progenitors, which are subsequently committed to innate lymphoid lineages in the α-lymphoid progenitor, early innate lymphoid progenitor, common helper innate lymphoid progenitor and innate lymphoid cell progenitor compartments. ILCs consist of conventional natural killer cells and helper-like cells (ILC1, ILC2 and ILC3). Despite recent advances, the cellular heterogeneity, developmental trajectory and signalling dependence of ILC progenitors are not fully understood. Here, using single-cell RNA-sequencing (scRNA-seq) of mouse bone marrow progenitors, we reveal ILC precursor subsets, delineate distinct ILC development stages and pathways, and report that high expression of programmed death 1 (PD-1hi) marked a committed ILC progenitor that was essentially identical to an innate lymphoid cell progenitor. Our data defined PD-1hiIL-25Rhi as an early checkpoint in ILC2 development, which was abolished by deficiency in the zinc-finger protein Bcl11b but restored by IL-25R overexpression. Similar to T lymphocytes, PD-1 was upregulated on activated ILCs. Administration of a PD-1 antibody depleted PD-1hi ILCs and reduced cytokine levels in an influenza infection model in mice, and blocked papain-induced acute lung inflammation. These results provide a perspective for exploring PD-1 and its ligand (PD-L1) in immunotherapy, and allow effective manipulation of the immune system for disease prevention and therapy.


Assuntos
Sequência de Bases , Linhagem da Célula , Imunidade Inata , Linfócitos/citologia , Células Progenitoras Linfoides/citologia , Receptor de Morte Celular Programada 1/metabolismo , Análise de Célula Única , Animais , Anticorpos/imunologia , Diferenciação Celular , Linhagem da Célula/genética , Separação Celular , Citocinas/imunologia , Citocinas/metabolismo , Modelos Animais de Doenças , Humanos , Imunoterapia/tendências , Influenza Humana/imunologia , Influenza Humana/metabolismo , Células Matadoras Naturais/citologia , Ativação Linfocitária , Linfócitos/imunologia , Linfócitos/metabolismo , Células Progenitoras Linfoides/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Pneumonia/imunologia , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Receptor de Morte Celular Programada 1/genética , Receptor de Morte Celular Programada 1/imunologia , Receptores de Interleucina/metabolismo , Proteínas Repressoras/deficiência , Proteínas Repressoras/metabolismo , Linfócitos T/metabolismo , Proteínas Supressoras de Tumor/deficiência , Proteínas Supressoras de Tumor/metabolismo
14.
Br J Nurs ; 31(17): 880-885, 2022 Sep 22.
Artigo em Inglês | MEDLINE | ID: mdl-36149420

RESUMO

Bloodstream infections associated with vascular access procedures pose a serious risk to patients that can be reduced by better standards of aseptic technique. The objectives of this roundtable of experts were to achieve a consensus on how to improve skin antisepsis in hospital, improve training, competency, compliance and consistency in skin antisepsis, review the role of devices in improving skin antisepsis, identify methods to improve skin antisepsis integrated with the Aseptic Non Touch Technique (ANTT®) approach, and identify challenges to the implementation of the panel's recommendations. Recommendations include using MHRA-licensed 2% chlorhexidine gluconate in 70% isopropyl alcohol solution with bidirectional strokes for up to 30 seconds, then leaving the skin to air dry for 30 seconds; using the ANTT Clinical Practice Framework and terminology as the standard for skin antisepsis training and practice; standardised ANTT and skin antisepsis education with 3-yearly competency assessments for all UK health professionals; and more research to address the evidence gap on transmission of infection after skin antisepsis.


Assuntos
Anti-Infecciosos Locais , Sepse , 2-Propanol , Anti-Infecciosos Locais/uso terapêutico , Antissepsia/métodos , Clorexidina/uso terapêutico , Humanos , Infecção da Ferida Cirúrgica
15.
J Virol ; 94(9)2020 04 16.
Artigo em Inglês | MEDLINE | ID: mdl-32075938

RESUMO

Recognition of influenza A virus (IAV) by the innate immune system triggers pathways that restrict viral replication, activate innate immune cells, and regulate adaptive immunity. However, excessive innate immune activation can exaggerate disease. The pathways promoting excessive activation are incompletely understood, with limited experimental models to investigate the mechanisms driving influenza virus-induced inflammation in humans. Interferon regulatory factor 5 (IRF5) is a transcription factor that plays important roles in the induction of cytokines after viral sensing. In an in vivo model of IAV infection, IRF5 deficiency reduced IAV-driven immune pathology and associated inflammatory cytokine production, specifically reducing cytokine-producing myeloid cell populations in Irf5-/- mice but not impacting type 1 interferon (IFN) production or virus replication. Using cytometry by time of flight (CyTOF), we identified that human lung IRF5 expression was highest in cells of the myeloid lineage. To investigate the role of IRF5 in mediating human inflammatory responses by myeloid cells to IAV, we employed human-induced pluripotent stem cells (hIPSCs) with biallelic mutations in IRF5, demonstrating for the first time that induced pluripotent stem cell-derived dendritic cells (iPS-DCs) with biallelic mutations can be used to investigate the regulation of human virus-induced immune responses. Using this technology, we reveal that IRF5 deficiency in human DCs, or macrophages, corresponded with reduced virus-induced inflammatory cytokine production, with IRF5 acting downstream of Toll-like receptor 7 (TLR7) and, possibly, retinoic acid-inducible gene I (RIG-I) after viral sensing. Thus, IRF5 acts as a regulator of myeloid cell inflammatory cytokine production during IAV infection in mice and humans and drives immune-mediated viral pathogenesis independently of type 1 IFN and virus replication.IMPORTANCE The inflammatory response to influenza A virus (IAV) participates in infection control but contributes to disease severity. After viral detection, intracellular pathways are activated, initiating cytokine production, but these pathways are incompletely understood. We show that interferon regulatory factor 5 (IRF5) mediates IAV-induced inflammation and, in mice, drives pathology. This was independent of antiviral type 1 IFN and virus replication, implying that IRF5 could be specifically targeted to treat influenza virus-induced inflammation. We show for the first time that human iPSC technology can be exploited in genetic studies of virus-induced immune responses. Using this technology, we deleted IRF5 in human myeloid cells. These IRF5-deficient cells exhibited impaired influenza virus-induced cytokine production and revealed that IRF5 acts downstream of Toll-like receptor 7 and possibly retinoic acid-inducible gene I. Our data demonstrate the importance of IRF5 in influenza virus-induced inflammation, suggesting that genetic variation in the IRF5 gene may influence host susceptibility to viral diseases.


Assuntos
Células-Tronco Pluripotentes Induzidas/imunologia , Vírus da Influenza A/imunologia , Fatores Reguladores de Interferon/metabolismo , Imunidade Adaptativa/fisiologia , Animais , Modelos Animais de Doenças , Interações Hospedeiro-Patógeno/imunologia , Humanos , Imunidade Inata/fisiologia , Vírus da Influenza A/metabolismo , Vírus da Influenza A/fisiologia , Influenza Humana/imunologia , Fatores Reguladores de Interferon/imunologia , Interferon Tipo I/metabolismo , Pulmão/virologia , Macrófagos/virologia , Camundongos , Infecções por Orthomyxoviridae/virologia , Replicação Viral/fisiologia
16.
Proc Natl Acad Sci U S A ; 115(41): 10428-10433, 2018 10 09.
Artigo em Inglês | MEDLINE | ID: mdl-30262653

RESUMO

Nontyphoidal Salmonellae cause a devastating burden of invasive disease in sub-Saharan Africa with high levels of antimicrobial resistance. Vaccination has potential for a major global health impact, but no licensed vaccine is available. The lack of commercial incentive makes simple, affordable technologies the preferred route for vaccine development. Here we compare equivalent Generalized Modules for Membrane Antigens (GMMA) outer membrane vesicles and O-antigen-CRM197 glycoconjugates to deliver lipopolysaccharide O-antigen in bivalent Salmonella Typhimurium and Enteritidis vaccines. Salmonella strains were chosen and tolR deleted to induce GMMA production. O-antigens were extracted from wild-type bacteria and conjugated to CRM197 Purified GMMA and glycoconjugates were characterized and tested in mice for immunogenicity and ability to reduce Salmonella infection. GMMA and glycoconjugate O-antigen had similar structural characteristics, O-acetylation, and glucosylation levels. Immunization with GMMA induced higher anti-O-antigen IgG than glycoconjugate administered without Alhydrogel adjuvant. With Alhydrogel, antibody levels were similar. GMMA induced a diverse antibody isotype profile with greater serum bactericidal activity than glycoconjugate, which induced almost exclusively IgG1. Immunization reduced bacterial colonization of mice subsequently infected with SalmonellaS Typhimurium numbers were lower in tissues of mice vaccinated with GMMA compared with glycoconjugate. S. Enteritidis burden in the tissues was similar in mice immunized with either vaccine. With favorable immunogenicity, low cost, and ability to induce functional antibodies and reduce bacterial burden, GMMA offer a promising strategy for the development of a nontyphoidal Salmonella vaccine compared with established glycoconjugates. GMMA technology is potentially attractive for development of vaccines against other bacteria of global health significance.


Assuntos
Anticorpos Antibacterianos/imunologia , Glicoconjugados/imunologia , Antígenos O/imunologia , Infecções por Salmonella/imunologia , Vacinas contra Salmonella/uso terapêutico , Salmonella enteritidis/imunologia , Salmonella typhimurium/imunologia , Animais , Anticorpos Antibacterianos/sangue , Camundongos , Infecções por Salmonella/prevenção & controle , Vacinação
17.
Proc Natl Acad Sci U S A ; 115(40): 10118-10123, 2018 10 02.
Artigo em Inglês | MEDLINE | ID: mdl-30217896

RESUMO

Intestinal epithelial cells (IECs) play a key role in regulating immune responses and controlling infection. However, the direct role of IECs in restricting pathogens remains incompletely understood. Here, we provide evidence that IL-22 primed intestinal organoids derived from healthy human induced pluripotent stem cells (hIPSCs) to restrict Salmonella enterica serovar Typhimurium SL1344 infection. A combination of transcriptomics, bacterial invasion assays, and imaging suggests that IL-22-induced antimicrobial activity is driven by increased phagolysosomal fusion in IL-22-pretreated cells. The antimicrobial phenotype was absent in hIPSCs derived from a patient harboring a homozygous mutation in the IL10RB gene that inactivates the IL-22 receptor but was restored by genetically complementing the IL10RB deficiency. This study highlights a mechanism through which the IL-22 pathway facilitates the human intestinal epithelium to control microbial infection.


Assuntos
Células Epiteliais/imunologia , Células-Tronco Pluripotentes Induzidas/imunologia , Interleucinas/imunologia , Mucosa Intestinal/imunologia , Fagossomos/imunologia , Infecções por Salmonella/imunologia , Salmonella typhimurium/imunologia , Células Epiteliais/microbiologia , Células Epiteliais/patologia , Humanos , Células-Tronco Pluripotentes Induzidas/microbiologia , Células-Tronco Pluripotentes Induzidas/patologia , Subunidade beta de Receptor de Interleucina-10/genética , Subunidade beta de Receptor de Interleucina-10/imunologia , Subunidade alfa de Receptor de Interleucina-21/genética , Subunidade alfa de Receptor de Interleucina-21/imunologia , Interleucinas/genética , Mucosa Intestinal/microbiologia , Mucosa Intestinal/patologia , Fagossomos/genética , Fagossomos/microbiologia , Fagossomos/patologia , Infecções por Salmonella/genética , Infecções por Salmonella/patologia , Salmonella typhimurium/genética , Interleucina 22
18.
Br J Nurs ; 30(1): 8-14, 2021 Jan 14.
Artigo em Inglês | MEDLINE | ID: mdl-33433273

RESUMO

This article discusses the importance of effective skin antisepsis prior to the insertion of peripheral intravenous catheters (PIVCs) and how best clinical practice is promoted by application of an appropriate method of skin disinfection integrated effectively with a proprietary aseptic non touch technique, or other standard aseptic technique. Historically under-reported, incidence of infection and risk to patients from PIVCs is now increasingly being recognised, with new research and evidence raising concern and helping to drive new clinical guidance and improvement. The risks posed by PIVCs are particularly significant given increasing PIVC dwell times, due to cannula removal now being determined by new guidance for clinical indication, rather than predefined time frames. Clinical 'best practice' is considered in context of the evidence base, importantly including availability and access to appropriate skin antisepsis products. In the UK, and other countries, ChloraPrep is the only skin antisepsis applicator licensed as a drug to disinfect skin and help prevent infections before invasive medical procedures, such as injections, blood sampling, insertion of PIVCs and minor or major surgery.


Assuntos
Antissepsia , Cateterismo Periférico , Pele , Antissepsia/métodos , Cateterismo Periférico/enfermagem , Enfermagem Baseada em Evidências , Humanos
19.
Gene Ther ; 27(9): 459-469, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32533104

RESUMO

Chronic granulomatous disease (CGD) is an inherited primary immunodeficiency disorder characterised by recurrent and often life-threatening infections and hyperinflammation. It is caused by defects of the phagocytic NADPH oxidase, a multicomponent enzyme system responsible for effective pathogen killing. A phase I/II clinical trial of lentiviral gene therapy is underway for the most common form of CGD, X-linked, caused by mutations in the gp91phox subunit of the NADPH oxidase. We propose to use a similar strategy to tackle p47phox-deficient CGD, caused by mutations in NCF1, which encodes the p47phox cytosolic component of the enzymatic complex. We generated a pCCLCHIM-p47phox lentiviral vector, containing the chimeric Cathepsin G/FES myeloid promoter and a codon-optimised version of the human NCF1 cDNA. Here we show that transduction with the pCCLCHIM-p47phox vector efficiently restores p47phox expression and biochemical NADPH oxidase function in p47phox-deficient human and murine cells. We also tested the ability of our gene therapy approach to control infection by challenging p47phox-null mice with Salmonella Typhimurium, a leading cause of sepsis in CGD patients, and found that mice reconstituted with lentivirus-transduced hematopoietic stem cells had a reduced bacterial load compared with untreated mice. Overall, our results potentially support the clinical development of a gene therapy approach using the pCCLCHIM-p47phox vector.


Assuntos
Doença Granulomatosa Crônica , Infecções por Salmonella , Animais , Humanos , Camundongos , Terapia Genética , Doença Granulomatosa Crônica/genética , Doença Granulomatosa Crônica/terapia , NADPH Oxidases/genética
20.
Immunology ; 159(4): 393-403, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-31880316

RESUMO

Signalling lymphocyte activation molecule family member 9 (SLAMF9) is an orphan receptor of the CD2/SLAM family of leucocyte surface proteins. Examination of SLAMF9 expression and function indicates that SLAMF9 promotes inflammation by specialized subsets of antigen-presenting cells. Within healthy liver and circulating mouse peripheral blood mononuclear cells, SLAMF9 is expressed on CD11b+ , Ly6C- , CD11clow , F4/80low , MHC-II+ , CX3 CR1+ mononuclear phagocytes as well as plasmacytoid dendritic cells. In addition, SLAMF9 can be found on peritoneal B1 cells and small (F4/80low ), but not large (F4/80high ), peritoneal macrophages. Upon systemic challenge with Salmonella enterica Typhimurium, Slamf9-/- mice were impaired in their ability to clear the infection from the liver. In humans, SLAMF9 is up-regulated upon differentiation of monocytes into macrophages, and lipopolysaccharide stimulation of PMA-differentiated, SLAMF9 knockdown THP-1 cells showed an essential role of SLAMF9 in production of granulocyte-macrophage colony-stimulating factor, tumour necrosis factor-α, and interleukin-1ß. Taken together, these data implicate SLAMF9 in the initiation of inflammation and clearance of bacterial infection.


Assuntos
Células Dendríticas/imunologia , Interações Hospedeiro-Patógeno/imunologia , Fígado/imunologia , Infecções por Salmonella/imunologia , Salmonella typhimurium/imunologia , Família de Moléculas de Sinalização da Ativação Linfocitária/imunologia , Animais , Linfócitos B/imunologia , Linfócitos B/microbiologia , Diferenciação Celular , Células Dendríticas/efeitos dos fármacos , Células Dendríticas/microbiologia , Regulação da Expressão Gênica , Fator Estimulador de Colônias de Granulócitos e Macrófagos/genética , Fator Estimulador de Colônias de Granulócitos e Macrófagos/imunologia , Interações Hospedeiro-Patógeno/genética , Humanos , Interleucina-1beta/genética , Interleucina-1beta/imunologia , Lipopolissacarídeos/farmacologia , Fígado/microbiologia , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Macrófagos/microbiologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Monócitos/efeitos dos fármacos , Monócitos/imunologia , Monócitos/microbiologia , Infecções por Salmonella/genética , Infecções por Salmonella/microbiologia , Salmonella typhimurium/patogenicidade , Transdução de Sinais , Família de Moléculas de Sinalização da Ativação Linfocitária/deficiência , Família de Moléculas de Sinalização da Ativação Linfocitária/genética , Células THP-1 , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/imunologia
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