Our studies in children with rheumatic diseases have led to the identification of two of the oldest cytokines, type I interferon (IFN) and interleukin 1 (IL-1), as important pathogenic players in ...systemic lupus erythematosus (SLE) and systemic onset juvenile arthritis (SoJIA), respectively. These findings were obtained by studying the transcriptional profiles of patient blood cells and by assessing the biological and transcriptional effect(s) of active patient sera on healthy blood cells. We also identified a signature that can be used to promptly diagnose SoJIA from other febrile conditions. Finally, our pilot clinical trials using IL-1 blockers have shown remarkable clinical benefits in SoJIA patients refractory to other medications.
As the capacity for generating large-scale molecular profiling data continues to grow, the ability to extract meaningful biological knowledge from it remains a limitation. Here, we describe the ...development of a new fixed repertoire of transcriptional modules, BloodGen3, that is designed to serve as a stable reusable framework for the analysis and interpretation of blood transcriptome data. The construction of this repertoire is based on co-clustering patterns observed across sixteen immunological and physiological states encompassing 985 blood transcriptome profiles. Interpretation is supported by customized resources, including module-level analysis workflows, fingerprint grid plot visualizations, interactive web applications and an extensive annotation framework comprising functional profiling reports and reference transcriptional profiles. Taken together, this well-characterized and well-supported transcriptional module repertoire can be employed for the interpretation and benchmarking of blood transcriptome profiles within and across patient cohorts. Blood transcriptome fingerprints for the 16 reference cohorts can be accessed interactively via: https://drinchai.shinyapps.io/BloodGen3Module/ .
The etiology of sporadic human chronic inflammatory diseases remains mostly unknown. To fill this gap, we developed a strategy that simultaneously integrates blood leukocyte responses to innate ...stimuli at the transcriptional, cellular, and secreted protein levels. When applied to systemic juvenile idiopathic arthritis (sJIA), an autoinflammatory disease of unknown etiology, this approach identified gene sets associated with specific cytokine environments and activated leukocyte subsets. During disease remission and off treatment, sJIA patients displayed dysregulated responses to TLR4, TLR8, and TLR7 stimulation. Isolated sJIA monocytes underexpressed the IL-1 inhibitor aryl hydrocarbon receptor (AHR) at baseline and accumulated higher levels of intracellular IL-1β after stimulation. Supporting the demonstration that AHR down-regulation skews monocytes toward macrophage differentiation, sJIA monocytes differentiated in vitro toward macrophages, away from the dendritic cell phenotype. This might contribute to the increased incidence of macrophage activation syndrome in these patients. Integrated analysis of high-dimensional data can thus unravel immune alterations predisposing to complex inflammatory diseases.
The development of novel human vaccines would be greatly facilitated by the development of in vivo models that permit preclinical analysis of human immune responses. Here, we show that nonobese ...diabetic severe combined immunodeficiency (NOD/SCID) β2 microglobulin−/− mice, engrafted with human CD34+ hematopoietic progenitors and further reconstituted with T cells, can mount specific immune responses against influenza virus vaccines. Live attenuated trivalent influenza virus vaccine induces expansion of CD8+ T cells specific to influenza matrix protein (FluM1) and nonstructural protein 1 in blood, spleen, and lungs. On ex vivo exposure to influenza antigens, antigen-specific CD8+ T cells produce IFN-γ and express cell-surface CD107a. FluM1-specific CD8+ T cells can be also expanded in mice vaccinated with inactivated trivalent influenza virus vaccine. Expansion of antigen-specific CD8+ T cells is dependent on reconstitution of the human myeloid compartment. Thus, this humanized mouse model permits preclinical testing of vaccines designed to induce cellular immunity, including those against influenza virus. Furthermore, this work sets the stage for systematic analysis of the in vivo functions of human DCs. This, in turn, will allow a new approach to the rational design and preclinical testing of vaccines that cannot be tested in human volunteers.
Sensing Pathogens and Tuning Immune Responses Pulendran, Bali; Palucka, Karolina; Banchereau, Jacques
Science (American Association for the Advancement of Science),
07/2001, Letnik:
293, Številka:
5528
Journal Article
Recenzirano
The immune system is capable of making qualitatively distinct responses against different microbial infections, and recent advances are starting to reveal how it manages this complex task. An ...integral component of the immune system is a network of cells known as dendritic cells (DCs), which sense different microbial stimuli and convey this information to lymphocytes. A better understanding of DC biology has allowed a model to be constructed in which the type of immune response to an infection is viewed as a function of several determinants, including the subpopulation of DCs, the nature of the microbe, microbe recognition receptors, and the cytokine microenvironment.
Monocytes differentiate into dendritic cells (DC) in response to GM‐CSF combined with other cytokines including IL‐4 and IL‐15. Here, we show that IL15‐DC are efficient in priming naive CD8+ T cells ...to differentiate into melanoma antigen‐specific cytotoxic T lymphocytes (CTL). While both melanoma peptide‐pulsed IL15‐DC and IL4‐DC expand high‐precursor frequency MART‐1‐specific CD8+ T cells after two stimulations in vitro, IL15‐DC require much lower peptide concentration for priming. IL15‐DC are more efficient in expanding gp100‐specific CD8+ T cells and can expand CD8+ T cells specific for Tyrosinase and MAGE‐3. CTL primed by IL15‐DC are superior in their function as demonstrated by (i) higher IFN‐γ secretion, (ii) higher expression of Granzyme B and Perforin, and (iii) higher killing of allogeneic melanoma cell lines, most particularly the HLA‐A*0201+ Sk‐Mel‐24 melanoma cells that are resistant to killing by CD8+ T cells primed with IL4‐DC. Supernatants of the sonicated cells demonstrate unique expression of IL‐1, IL‐8 and IL‐15. Therefore, membrane‐bound IL‐15 might contribute to enhanced priming by IL15‐DC. Thus, IL‐15 induces myeloid DC that are efficient in priming and maturation of melanoma antigen‐specific CTL.
Immunity results from a complex interplay between the antigen‐non‐specific innate immune system and the antigen‐specific adaptive immune system. The cells and molecules of the innate system employ ...non‐clonal recognition receptors including lectins, Toll‐like receptors, NOD‐like receptors, and helicases. B and T lymphocytes of the adaptive immune system employ clonal receptors recognizing antigens or their derived peptides in a highly specific manner. An essential link between innate and adaptive immunity is provided by dendritic cells (DCs). DCs can induce such contrasting states as immunity and tolerance. The recent years have brought a wealth of information on the biology of DCs revealing the complexity of this cell system. Indeed, DC plasticity and subsets are prominent determinants of the type and quality of elicited immune responses. In this article, we summarize our recent studies aimed at a better understanding of the DC system to unravel the pathophysiology of human diseases and design novel human vaccines.
B and T lymphocytes recognize antigens with high specificity, but neither initiate immune responses, nor decide their types. These functions rest upon dendritic cells (DCs), which can determine and ...maintain Th1/Th2 polarization. Immune responses are thus dependent on the DC subset, the receptors that recognize each pathogen and the microenvironment. Microbes employ an array of mechanisms to evade and disrupt DC functions; some even hijack DCs for transport around the body. Our progress in the understanding of DC physiology will hopefully help us create the necessary vaccines to counteract the infectious agents that still plague mankind.
Immune responses to microbes ultimately depend on the way that dendritic cells recognize and respond initially.
Background Respiratory syncytial virus (RSV) is the principal etiologic agent of bronchiolitis and viral pneumonia in infants and young children. Yet, many aspects of its immunopathogenesis are not ...well understood Methods We analyzed the immune cells that are mobilized by RSV and other respiratory viruses, by studying nasal wash samples from children hospitalized with acute viral respiratory infections Results RSV mobilizes virtually all blood immune cells, including myeloid dendritic cells (DCs) and plasmacytoid DCs (pDCs), to the nasal mucosa. DCs were also mobilized to the nasal mucosa of children with other viral respiratory infections. The increased number of pDCs in the nasal compartment significantly correlates with RSV load (P=.022), and it is associated with a significant decrease in the number of pDCs in the blood (P=.007). The influx of DCs in the nasal mucosa is not transient, as even higher numbers of both DC subsets were found in respiratory secretions weeks after the acute symptoms of RSV infection had resolved. Immunochemistry analysis of respiratory samples has demonstrated the presence of the RSV fusion protein within HLA-DR–positive cells Conclusion Infection with RSV and other respiratory viruses mobilizes DCs to the site of viral entry
Humanized mice, defined as mice with human immune systems, have become an emerging model to study human hematopoiesis, infectious disease, and cancer. Here, we describe the techniques to generate ...humanized NSGF6 mice using adult human CD34+ hematopoietic stem and progenitor cells (HSPCs). We describe steps for constructing and monitoring the engraftment of humanized mice. We then detail procedures for tissue processing and immunophenotyping by flow cytometry to evaluate the multilineage hematopoietic differentiation.
For complete details on the use and execution of this protocol, please refer to Yu et al.1
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•Humanized mouse model with murine Flt3 KO and human IL6 KI by adult HSPCs•Detailed steps for the construction of humanized mice•Techniques to monitor human engraftment in the blood•Procedures for tissue processing and immunophenotyping by flow cytometry
Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.
Humanized mice, defined as mice with human immune systems, have become an emerging model to study human hematopoiesis, infectious disease, and cancer. Here, we describe the techniques to generate humanized NSGF6 mice using adult human CD34+ hematopoietic stem and progenitor cells (HSPCs). We describe steps for constructing and monitoring the engraftment of humanized mice. We then detail procedures for tissue processing and immunophenotyping by flow cytometry to evaluate the multilineage hematopoietic differentiation.
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