RESUMO
During primary varicella zoster virus (VZV) infection, infected lymphocytes drive primary viremia, causing systemic dissemination throughout the host, including the skin. This results in cytokine expression, including interferons (IFNs), which partly limit infection. VZV also spreads from skin keratinocytes to lymphocytes prior to secondary viremia. It is not clear how VZV achieves this while evading the cytokine response. Here, we show that VZV glycoprotein C (gC) binds IFN-γ and modifies its activity, increasing the expression of a subset of IFN-stimulated genes (ISGs), including intercellular adhesion molecule 1 (ICAM1), chemokines and immunomodulatory genes. The higher ICAM1 protein level at the plasma membrane of keratinocytes facilitates lymphocyte function-associated antigen 1-dependent T cell adhesion and expression of gC during infection increases VZV spread to peripheral blood mononuclear cells. This constitutes the discovery of a strategy to modulate IFN-γ activity, upregulating a subset of ISGs, promoting enhanced lymphocyte adhesion and virus spread.
Assuntos
Adesão Celular , Herpesvirus Humano 3 , Molécula 1 de Adesão Intercelular , Interferon gama , Queratinócitos , Linfócitos T , Humanos , Interferon gama/metabolismo , Interferon gama/imunologia , Linfócitos T/imunologia , Linfócitos T/metabolismo , Linfócitos T/virologia , Molécula 1 de Adesão Intercelular/metabolismo , Molécula 1 de Adesão Intercelular/genética , Queratinócitos/virologia , Queratinócitos/metabolismo , Queratinócitos/imunologia , Herpesvirus Humano 3/fisiologia , Infecção pelo Vírus da Varicela-Zoster/imunologia , Infecção pelo Vírus da Varicela-Zoster/virologia , Leucócitos Mononucleares/virologia , Leucócitos Mononucleares/metabolismo , Leucócitos Mononucleares/imunologia , Proteínas do Envelope Viral/metabolismo , Antígeno-1 Associado à Função Linfocitária/metabolismoRESUMO
Systemic Lupus Erythematosus (SLE) is an autoimmune disease, the pathophysiology and genetic basis of which are incompletely understood. Using a forward genetic screen in multiplex families with systemic lupus erythematosus (SLE) we identified an association between SLE and compound heterozygous deleterious variants in the non-receptor tyrosine kinases (NRTKs) ACK1 and BRK. Experimental blockade of ACK1 or BRK increased circulating autoantibodies in vivo in mice and exacerbated glomerular IgG deposits in an SLE mouse model. Mechanistically, non-receptor tyrosine kinases (NRTKs) regulate activation, migration, and proliferation of immune cells. We found that the patients' ACK1 and BRK variants impair efferocytosis, the MERTK-mediated anti-inflammatory response to apoptotic cells, in human induced Pluripotent Stem Cell (hiPSC)-derived macrophages, which may contribute to SLE pathogenesis. Overall, our data suggest that ACK1 and BRK deficiencies are associated with human SLE and impair efferocytosis in macrophages.
RESUMO
Chimeric antigen receptor (CAR) macrophages have broadened the landscape of anti-cancer immunotherapies to combat solid malignancies. Shah et al. introduce CARs to facilitate a CAR macrophage therapy, which aims to recruit and activate T/natural killer cells, further strengthening the overall immune response to decrease pancreatic cancer burden and metastatic spreading.
Assuntos
Macrófagos , Receptores de Antígenos Quiméricos , Humanos , Macrófagos/imunologia , Receptores de Antígenos Quiméricos/imunologia , Neoplasias/imunologia , Neoplasias/terapia , Imunoterapia/métodos , Animais , Imunoterapia Adotiva/métodos , Células Matadoras Naturais/imunologia , Neoplasias Pancreáticas/imunologia , Neoplasias Pancreáticas/terapia , Neoplasias Pancreáticas/patologiaRESUMO
The Cystic Fibrosis Conductance Transmembrane Regulator gene encodes for the CFTR ion channel, which is responsible for the transport of chloride and bicarbonate across the plasma membrane. Mutations in the gene result in impaired ion transport, subsequently leading to perturbed secretion in all exocrine glands and, therefore, the multi-organ disease cystic fibrosis (CF). In recent years, several studies have reported on CFTR expression in immune cells as demonstrated by immunofluorescence, flow cytometry, and immunoblotting. However, these data are mainly restricted to single-cell populations and show significant variation depending on the methodology used. Here, we investigated CFTR transcription and protein expression using standardized protocols in a comprehensive panel of immune cells. Methods: We applied a high-resolution Western blot protocol using a combination of highly specific monoclonal CFTR antibodies that have been optimized for the detection of CFTR in epithelial cells and healthy primary immune cell subpopulations sorted by flow cytometry and used immortalized cell lines as controls. The specificity of CFTR protein detection was controlled by peptide competition and enzymatic Peptide-N-Glycosidase-F (PNGase) digest. CFTR transcripts were analyzed using quantitative real-time PCR and normalized to the level of epithelial T84 cells as a reference. Results: CFTR mRNA expression could be shown for primary CD4+ T cells, NK cells, as well as differentiated THP-1 and Jurkat T cells. In contrast, we failed to detect CFTR transcripts for CD14+ monocytes and undifferentiated THP-1 cells, as well as for B cells and CD8+ T cells. Prominent immunoreactive bands were detectable by immunoblotting with the combination of four CFTR antibodies targeting different epitopes of the CFTR protein. However, in biosamples of non-epithelial origin, these CFTR-like protein bands could be unmasked as false positives through peptide competition or PNGase digest, meaning that the observed mRNA transcripts were not necessarily translated into CFTR proteins, which could be detected via immunoblotting. Our results confirm that mRNA expression in immune cells is many times lower than in that cells of epithelial origin. The immunoreactive signals in immune cells turned out to be false positives, and may be provoked by the presence of a high-affinity protein with a similar epitope. Non-specific binding (e.g., Fab-interaction with glycosyl branches) might also contribute to false positive signals. Our findings highlight the necessity of accurate controls, such as CFTR-negative cells, as well as peptide competition and glycolytic digest in order to identify genuine CFTR protein by immunoblotting. Our data suggest, furthermore, that CFTR protein expression data from techniques such as histology, for which the absence of a molecular weight or other independent control prevents the unmasking of false positive immunoreactive signals, must be interpreted carefully as well.
Assuntos
Regulador de Condutância Transmembrana em Fibrose Cística , Leucócitos Mononucleares , RNA Mensageiro , Regulador de Condutância Transmembrana em Fibrose Cística/genética , Regulador de Condutância Transmembrana em Fibrose Cística/metabolismo , Humanos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Leucócitos Mononucleares/metabolismo , Western Blotting , Reação em Cadeia da Polimerase em Tempo Real/métodos , Fibrose Cística/metabolismo , Fibrose Cística/genética , Células Matadoras Naturais/metabolismo , Citometria de Fluxo/métodos , Linfócitos T CD4-Positivos/metabolismoRESUMO
BACKGROUND: There is a significant demand for intermediate-scale bioreactors in academic and industrial institutions to produce cells for various applications in drug screening and/or cell therapy. However, the application of these bioreactors in cultivating hiPSC-derived immune cells and other blood cells is noticeably lacking. To address this gap, we have developed a xeno-free and chemically defined intermediate-scale bioreactor platform, which allows for the generation of standardized human iPSC-derived hematopoietic organoids and subsequent continuous production of macrophages (iPSC-Mac). METHODS: We describe a novel method for intermediate-scale immune cell manufacturing, specifically the continuous production of functionally and phenotypically relevant macrophages that are harvested on weekly basis for multiple weeks. RESULTS: The continuous production of standardized human iPSC-derived macrophages (iPSC-Mac) from 3D hematopoietic organoids also termed hemanoids, is demonstrated. The hemanoids exhibit successive stage-specific embryonic development, recapitulating embryonic hematopoiesis. iPSC-Mac were efficiently and continuously produced from three different iPSC lines and exhibited a consistent and reproducible phenotype, as well as classical functionality and the ability to adapt towards pro- and anti-inflammatory activation stages. Single-cell transcriptomic analysis revealed high macrophage purity. Additionally, we show the ability to use the produced iPSC-Mac as a model for testing immunomodulatory drugs, exemplified by dexamethasone. CONCLUSIONS: The novel method demonstrates an easy-to-use intermediate-scale bioreactor platform that produces prime macrophages from human iPSCs. These macrophages are functionally active and require no downstream maturation steps, rendering them highly desirable for both therapeutic and non-therapeutic applications.
Assuntos
Reatores Biológicos , Células-Tronco Pluripotentes Induzidas , Macrófagos , Organoides , Humanos , Células-Tronco Pluripotentes Induzidas/citologia , Células-Tronco Pluripotentes Induzidas/metabolismo , Macrófagos/citologia , Macrófagos/metabolismo , Organoides/citologia , Organoides/metabolismo , Diferenciação Celular , Técnicas de Cultura de Células/métodos , Técnicas de Cultura de Células/instrumentação , HematopoeseRESUMO
Ensuring the safety of parenteral drugs before injection into patients is of utmost importance. New regulations around the globe and the need to refrain from using animals however, have highlighted the need for new cell sources to be used in next-generation bioassays to detect the entire spectrum of possible contaminating pyrogens. Given the current drawbacks of the Monocyte-Activation-Test (MAT) with respect to the use of primary peripheral blood mono-nuclear cells or the use of monocytic cell lines, we here demonstrate the manufacturing of sensor monocytes/macrophages from human induced pluripotent stem cells (iMonoMac), which are fully defined and superior to current cell products. Using a modern and scalable manufacturing platform, iMonoMac showed typical macrophage-like morphology and stained positive for several Toll like receptor (TLRs) such as TLR-2, TLR-5, TLR-4. Furthermore, iMonoMac derived from the same donor were sensitive to endotoxins, non-endotoxins, and process related pyrogens at a high dynamic range and across different cellular densities. Of note, iMonoMac showed increased sensitivity and reactivity to a broad range of pyrogens, demonstrated by the detection of interleukin-6 at low concentrations of LPS and MALP-2 which could not be reached using the current MAT cell sources. To further advance the system, iMonoMac or genetically engineered iMonoMac with NF-κB-luciferase reporter cassette could reveal a specific activation response while correlating to the classical detection method employing enzyme-linked immunosorbent assay to measure cytokine secretion. Thus, we present a valuable cellular tool to assess parenteral drugs safety, facilitating the future acceptance and design of regulatory-approved bioassays.
Assuntos
Células-Tronco Pluripotentes Induzidas , Macrófagos , Pirogênios , Células-Tronco Pluripotentes Induzidas/citologia , Células-Tronco Pluripotentes Induzidas/metabolismo , Humanos , Macrófagos/metabolismo , Macrófagos/efeitos dos fármacos , Macrófagos/citologia , Contaminação de Medicamentos , Receptores Toll-Like/metabolismo , Endotoxinas , Interleucina-6/metabolismo , Monócitos/citologia , Monócitos/metabolismo , Monócitos/efeitos dos fármacos , Infusões ParenteraisRESUMO
Fabry disease (FD) is a rare and inherited monogenetic disease caused by mutations in the X-chromosomal alpha-galactosidase A gene GLA concomitant with accumulation of its substrate globotriaosylceramide (Gb3) and multi-organ symptoms. We derived an induced pluripotent stem cell line, MHHi029-A, from a male FD patient carrying a c.959A > T missense mutation in the GLA gene. The hiPSCs show a normal karyotype, expression of pluripotency markers and trilineage differentiation capacity. Importantly, they present the patient-specific mutation in the GLA gene and are therefore a valuable resource for investigating the FD mechanism and identifying novel therapies.
Assuntos
Doença de Fabry , Células-Tronco Pluripotentes Induzidas , alfa-Galactosidase , Doença de Fabry/genética , Doença de Fabry/patologia , Humanos , Células-Tronco Pluripotentes Induzidas/metabolismo , Masculino , alfa-Galactosidase/genética , alfa-Galactosidase/metabolismo , Linhagem Celular , Diferenciação Celular , MutaçãoRESUMO
The CRISPR-Cas12a platform has attracted interest in the genome editing community because the prototypical Acidaminococcus Cas12a generates a staggered DNA double-strand break upon binding to an AT-rich protospacer-adjacent motif (PAM, 5'-TTTV). The broad application of the platform in primary human cells was enabled by the development of an engineered version of the natural Cas12a protein, called Cas12a Ultra. In this study, we confirmed that CRISPR-Cas12a Ultra ribonucleoprotein complexes enabled allelic gene disruption frequencies of over 90% at multiple target sites in human T cells, hematopoietic stem and progenitor cells (HSPCs), and induced pluripotent stem cells (iPSCs). In addition, we demonstrated, for the first time, the efficient knock-in potential of the platform in human iPSCs and achieved targeted integration of a GFP marker gene into the AAVS1 safe harbor site and a CSF2RA super-exon into CSF2RA in up to 90% of alleles without selection. Clonal analysis revealed bi-allelic integration in >50% of the screened iPSC clones without compromising their pluripotency and genomic integrity. Thus, in combination with the adeno-associated virus vector system, CRISPR-Cas12a Ultra provides a highly efficient genome editing platform for performing targeted knock-ins in human iPSCs.
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Células-Tronco Pluripotentes Induzidas , Células-Tronco Pluripotentes , Humanos , Sistemas CRISPR-Cas , Células-Tronco Hematopoéticas , AlelosRESUMO
Pulmonary alveolar proteinosis (PAP) is an umbrella term used to refer to a pulmonary syndrome which is characterized by excessive accumulation of surfactant in the lungs of affected individuals. In general, PAP is a rare lung disease affecting children and adults, although its prevalence and incidence is variable among different countries. Even though PAP is a rare disease, it is a prime example on how modern medicine can lead to new therapeutic concepts, changing ways and techniques of (genetic) diagnosis which ultimately led into personalized treatments, all dedicated to improve the function of the impaired lung and thus life expectancy and quality of life in PAP patients. In fact, new technologies, such as new sequencing technologies, gene therapy approaches, new kind and sources of stem cells and completely new insights into the ontogeny of immune cells such as macrophages have increased our understanding in the onset and progression of PAP, which have paved the way for novel therapeutic concepts for PAP and beyond. As of today, classical monocyte-derived macrophages are known as important immune mediator and immune sentinels within the innate immunity. Furthermore, macrophages (known as tissue resident macrophages (TRMs)) can also be found in various tissues, introducing e. g. alveolar macrophages in the broncho-alveolar space as crucial cellular determinants in the onset of PAP and other lung disorders. Given recent insights into the onset of alveolar macrophages and knowledge about factors which impede their function, has led to the development of new therapies, which are applied in the context of PAP, with promising implications also for other diseases in which macrophages play an important role. Thus, we here summarize the latest insights into the various forms of PAP and introduce new pre-clinical work which is currently conducted in the framework of PAP, introducing new therapies for children and adults who still suffer from this severe, potentially life-threatening disease.
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Proteinose Alveolar Pulmonar , Adulto , Criança , Humanos , Proteinose Alveolar Pulmonar/diagnóstico , Proteinose Alveolar Pulmonar/genética , Proteinose Alveolar Pulmonar/terapia , Qualidade de Vida , Pulmão , Macrófagos AlveolaresRESUMO
Alveolar macrophages (AMs) represent crucial immune cells in the bronchioalveolar space of the lung. Given the important role in the host defense machinery and lung tissue homeostasis, AMs have been linked to a variety of diseases and thus represent a promising target cell type for novel therapies. The emerging importance of AM underlines the necessity to isolate and/or generate proper cellular models, which facilitate basic biology and translational science. As of yet, most studies focus on the derivation of AM from the murine system. This chapter introduces the use of human-induced pluripotent stem cell (iPSC)-derived primitive macrophages, which can be further matured towards an AM-like phenotype upon intra-pulmonary transfer into mice. We will give a brief overview on the generation of primitive iPSC-derived macrophages, which is followed by a detailed, step-by-step description of the intra-pulmonary transfer of cells and the follow-up procedures needed to isolate the iPSC-derived, AM-like cells from the lungs post-transfer. The chapter provides an alternative approach to derive human AM-like cells, which can be used to study human AM biology and to investigate novel therapeutic interventions using primitive macrophages from iPSC.
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Células-Tronco Pluripotentes Induzidas , Macrófagos Alveolares , Humanos , Animais , Camundongos , Modelos Animais de Doenças , Macrófagos , FenótipoRESUMO
We describe a human lung disease caused by autosomal recessive, complete deficiency of the monocyte chemokine receptor C-C motif chemokine receptor 2 (CCR2). Nine children from five independent kindreds have pulmonary alveolar proteinosis (PAP), progressive polycystic lung disease, and recurrent infections, including bacillus Calmette Guérin (BCG) disease. The CCR2 variants are homozygous in six patients and compound heterozygous in three, and all are loss-of-expression and loss-of-function. They abolish CCR2-agonist chemokine C-C motif ligand 2 (CCL-2)-stimulated Ca2+ signaling in and migration of monocytic cells. All patients have high blood CCL-2 levels, providing a diagnostic test for screening children with unexplained lung or mycobacterial disease. Blood myeloid and lymphoid subsets and interferon (IFN)-γ- and granulocyte-macrophage colony-stimulating factor (GM-CSF)-mediated immunity are unaffected. CCR2-deficient monocytes and alveolar macrophage-like cells have normal gene expression profiles and functions. By contrast, alveolar macrophage counts are about half. Human complete CCR2 deficiency is a genetic etiology of PAP, polycystic lung disease, and recurrent infections caused by impaired CCL2-dependent monocyte migration to the lungs and infected tissues.
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Proteinose Alveolar Pulmonar , Receptores CCR2 , Criança , Humanos , Pulmão/metabolismo , Macrófagos Alveolares/metabolismo , Proteinose Alveolar Pulmonar/genética , Proteinose Alveolar Pulmonar/diagnóstico , Receptores CCR2/deficiência , Receptores CCR2/genética , Receptores CCR2/metabolismo , Reinfecção/metabolismoRESUMO
BACKGROUND: Interstitial lung diseases (ILD) are a group of rare lung diseases with severe outcomes. The COST Innovator Grant aims to establish a first-of-a-kind open-access Biorepository of patient-derived induced pluripotent stem cells (iPSC) and to train researchers in the skills required to generate a robust preclinical model of ILD using these cells. This study aims to describe and evaluate the effectiveness of a training course designed to train researchers in iPSC techniques to model ILD. METHODS: 74 researchers, physicians and stakeholders attended the training course in Dublin in May 2022 with 31 trainees receiving teaching in practical iPSC culturing skills. The training course learners were divided into the Hands-on (16 trainees) and Observer groups (15 trainees), with the Observers attending a supervised live-streamed experience of the laboratories skills directly delivered to the Hands-on group. All participants were asked to participate in an evaluation to analyse their satisfaction and knowledge gained during the Training Course, with means compared using t-tests. RESULTS: The gender balance in both groups was predominantly females (77.4%). The Hands-on group consisted mainly of researchers (75%), whereas all participants of the Observer group described themselves as clinicians. All participants in the Hands-on group were at least very satisfied with the training course compared to 70% of the participants in the Observer group. The knowledge assessment showed that the Hands-on group retained significantly more knowledge of iPSC characteristics and culturing techniques compared to the Observers (* < 0.05; p = 0.0457). A comprehensive learning video detailing iPSC culturing techniques was produced and is included with this manuscript. CONCLUSIONS: The majority of participants were highly or very satisfied with the training course and retained significant knowledge about iPSC characteristics and culturing techniques after attending the training course. Overall, our findings demonstrate the feasibility of running hybrid Hands-on and Observer teaching events and underscore the importance of this type of training programme to appeal to a broad spectrum of interested clinicians and researchers particularly in rare disease. The long-term implications of this type of training event requires further study to determine its efficacy and impact on adoption of iPSC disease modelling techniques in participants' laboratories.
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Células-Tronco Pluripotentes Induzidas , Doenças Pulmonares Intersticiais , Feminino , Humanos , Masculino , Acesso à Informação , Doenças Pulmonares Intersticiais/terapiaRESUMO
BACKGROUND: Macrophages have recently become attractive therapeutics in cancer immunotherapy. The potential of macrophages to infiltrate and influence solid malignancies makes them promising targets for the chimeric antigen receptor (CAR) technology to redirect their stage of polarization, thus enhancing their anticancer capacities. Given the emerging interest for CAR-macrophages, generation of such cells so far mainly depends on peripheral blood monocytes, which are isolated from the respective donor prior to genetic manipulation. This procedure is time-intensive and cost-intensive, while, in some cases, insufficient monocyte amounts can be recovered from the donor, thus hampering the broad applicability of this technology. Hence, we demonstrate the generation and effectiveness of CAR-macrophages from various stem cell sources using also modern upscaling technologies for next generation immune cell farming. METHODS: Primary human hematopoietic stem and progenitor cells and induced pluripotent stem cells were used to derive anti-CD19 CAR-macrophages. Anticancer activity of the cells was demonstrated in co-culture systems, including primary material from patients with leukemia. Generation of CAR-macrophages was facilitated by bioreactor technologies and single-cell RNA (scRNA) sequencing was used to characterize in-depth response and behavior of CAR-macrophages. RESULTS: Irrespective of the stem-cell source, CAR-macrophages exhibited enhanced and antigen-dependent phagocytosis of CD19+ target cancer cells with increased pro-inflammatory responses. Phagocytic capacity of CAR-macrophages was dependent on target cell CD19 expression levels with superior function of CAR-macrophages against CD19+ cancer cell lines and patient-derived acute lymphocytic leukemia cancer cells. scRNA sequencing revealed CAR-macrophages to be distinct from eGFP control cells after co-culture with target cells, which includes the activation of pro-inflammatory pathways and upregulation of chemokines and cytokines associated with adaptive immune cell recruitment, favoring the repolarization of CAR-macrophages to a pro-inflammatory state. Taken together, the data highlight the unique features of CAR-macrophages in combination with the successful upscaling of the production pipeline using a three-dimensional differentiation protocol and intermediate scale bioreactors. CONCLUSION: In summary, our work provides insights into the seminal use and behavior of CAR-macrophages which are derived from various sources of stem cells, while introducing a unique technology for CAR-macrophage manufacturing, all dedicated to the clinical translation of CAR-macrophages within the field of anticancer immunotherapies.
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Células-Tronco Pluripotentes Induzidas , Leucemia , Neoplasias , Receptores de Antígenos Quiméricos , Humanos , Receptores de Antígenos Quiméricos/genética , Receptores de Antígenos de Linfócitos T , Células-Tronco Pluripotentes Induzidas/metabolismo , Linfócitos T , Leucemia/terapia , Macrófagos/metabolismoRESUMO
Alveolar macrophages (AMs) are the sentinel cells of the alveolar space, maintaining homeostasis, fending off pathogens, and controlling lung inflammation. During acute lung injury, AMs orchestrate the initiation and resolution of inflammation in order to ultimately restore homeostasis. This central role in acute lung inflammation makes AMs attractive targets for therapeutic interventions. Single-cell RNA-Seq and spatial omics approaches, together with methodological advances such as the generation of human macrophages from pluripotent stem cells, have increased understanding of the ontogeny, function, and plasticity of AMs during infectious and sterile lung inflammation, which could move the field closer to clinical application. However, proresolution phenotypes might conflict with proinflammatory and antibacterial responses. Therefore, therapeutic targeting of AMs at vulnerable time points over the course of infectious lung injury might harbor the risk of serious side effects, such as loss of antibacterial host defense capacity. Thus, the identification of key signaling hubs that determine functional fate decisions in AMs is of the utmost importance to harness their therapeutic potential.
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Lesão Pulmonar Aguda , Pneumonia , Humanos , Macrófagos Alveolares , Inflamação , Homeostase , PulmãoRESUMO
Abnormalities at any stage of trophoblast development may result in pregnancy-related complications. Many of these adverse outcomes are discovered later in pregnancy, but the underlying pathomechanisms are constituted during the first trimester. Acquiring developmentally relevant material to elucidate the disease mechanisms is difficult. Human pluripotent stem cell (hPSC) technology can provide a renewable source of relevant cells. BMP4, A83-01, and PD173074 (BAP) treatment drives trophoblast commitment of hPSCs toward syncytiotrophoblast (STB), but lacks extravillous trophoblast (EVT) cells. EVTs mediate key functions during placentation, remodeling of uterine spiral arteries, and maintenance of immunological tolerance. We optimized the protocol for a more efficient generation of HLA-Gpos EVT-like trophoblasts from primed hiPSCs. Increasing the concentrations of A83-01 and PD173074, while decreasing bulk cell density resulted in an increase in HLA-G of up to 71%. Gene expression profiling supports the advancements of our treatment regarding the generation of trophoblast cells. The reported differentiation protocol will allow for an on-demand access to human trophoblast cells enriched for HLA-Gpos EVT-like cells, allowing for the elucidation of placenta-related disorders and investigating the immunological tolerance toward the fetus, overcoming the difficulties in obtaining primary EVTs without the need for a complex differentiation pathway via naïve pluripotent or trophoblast stem cells.
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Antígenos HLA-G , Células-Tronco Pluripotentes Induzidas , Feminino , Gravidez , Humanos , Trofoblastos , Diferenciação CelularRESUMO
The skin needs to balance tolerance of colonizing microflora with rapid detection of potential pathogens. Flexible response mechanisms would seem most suitable to accommodate the dynamic challenges of effective antimicrobial defense and restoration of tissue homeostasis. Here, we dissected macrophage-intrinsic mechanisms and microenvironmental cues that tune macrophage signaling in localized skin infection with the colonizing and opportunistic pathogen Staphylococcus aureus. Early in skin infection, the cytokine granulocyte-macrophage colony-stimulating factor (GM-CSF) produced by γδ T cells and hypoxic conditions within the dermal microenvironment diverted macrophages away from a homeostatic M-CSF- and hypoxia-inducible factor 1α (HIF-1α)-dependent program. This allowed macrophages to be metabolically rewired for maximal inflammatory activity, which requires expression of Irg1 and generation of itaconate, but not HIF-1α. This multifactorial macrophage rewiring program was required for both the timely clearance of bacteria and for the provision of local immune memory. These findings indicate that immunometabolic conditioning allows dermal macrophages to cycle between antimicrobial activity and protection against secondary infections.