A local regulatory T cell feedback circuit maintains immune homeostasis by pruning self-activated T cells.

A fraction of mature T cells can be activated by peripheral self-antigens, potentially eliciting host autoimmunity. We investigated homeostatic control of self-activated T cells within unperturbed tissue environments by combining high-resolution multiplexed and volumetric imaging with computational modeling. In lymph nodes, self-activated T cells produced interleukin (IL)-2, which enhanced local regulatory T cell (Treg) proliferation and inhibitory functionality. The resulting micro-domains reciprocally constrained inputs required for damaging effector responses, including CD28 co-stimulation and IL-2 signaling, constituting a negative feedback circuit. Due to these local constraints, self-activated T cells underwent transient clonal expansion, followed by rapid death ("pruning"). Computational simulations and experimental manipulations revealed the feedback machinery's quantitative limits: modest reductions in Treg micro-domain density or functionality produced non-linear breakdowns in control, enabling self-activated T cells to subvert pruning. This fine-tuned, paracrine feedback process not only enforces immune homeostasis but also establishes a sharp boundary between autoimmune and host-protective T cell responses.

Adaptive immune responses to SARS-CoV-2 persist in the pharyngeal lymphoid tissue of children.

Most studies of adaptive immunity to SARS-CoV-2 infection focus on peripheral blood, which may not fully reflect immune responses at the site of infection. Using samples from 110 children undergoing tonsillectomy and adenoidectomy during the COVID-19 pandemic, we identified 24 samples with evidence of previous SARS-CoV-2 infection, including neutralizing antibodies in serum and SARS-CoV-2-specific germinal center and memory B cells in the tonsils and adenoids. Single-cell B cell receptor (BCR) sequencing indicated virus-specific BCRs were class-switched and somatically hypermutated, with overlapping clones in the two tissues. Expanded T cell clonotypes were found in tonsils, adenoids and blood post-COVID-19, some with CDR3 sequences identical to previously reported SARS-CoV-2-reactive T cell receptors (TCRs). Pharyngeal tissues from COVID-19-convalescent children showed persistent expansion of germinal center and antiviral lymphocyte populations associated with interferon (IFN)-γ-type responses, particularly in the adenoids, and viral RNA in both tissues. Our results provide evidence for persistent tissue-specific immunity to SARS-CoV-2 in the upper respiratory tract of children after infection.

Resident Macrophages Cloak Tissue Microlesions to Prevent Neutrophil-Driven Inflammatory Damage.

Neutrophils are attracted to and generate dense swarms at sites of cell damage in diverse tissues, often extending the local disruption of organ architecture produced by the initial insult. Whether the inflammatory damage resulting from such neutrophil accumulation is an inescapable consequence of parenchymal cell death has not been explored. Using a combination of dynamic intravital imaging and confocal multiplex microscopy, we report here that tissue-resident macrophages rapidly sense the death of individual cells and extend membrane processes that sequester the damage, a process that prevents initiation of the feedforward chemoattractant signaling cascade that results in neutrophil swarms. Through this "cloaking" mechanism, the resident macrophages prevent neutrophil-mediated inflammatory damage, maintaining tissue homeostasis in the face of local cell injury that occurs on a regular basis in many organs because of mechanical and other stresses. VIDEO ABSTRACT.

Systems biology in immunology: a computational modeling perspective.

Systems biology is an emerging discipline that combines high-content, multiplexed measurements with informatic and computational modeling methods to better understand biological function at various scales. Here we present a detailed review of the methods used to create computational models and to conduct simulations of immune function. We provide descriptions of the key data-gathering techniques employed to generate the quantitative and qualitative data required for such modeling and simulation and summarize the progress to date in applying these tools and techniques to questions of immunological interest, including infectious disease. We include comments on what insights modeling can provide that complement information obtained from the more familiar experimental discovery methods used by most investigators and the reasons why quantitative methods are needed to eventually produce a better understanding of immune system operation in health and disease.

AI and immunology.

AI is rapidly becoming part of many aspects of daily life, with an impact that reaches all fields of research. We asked investigators to share their thoughts on how AI is changing immunology research, what is necessary to move forward, the potential and the pitfalls, and what will remain unchanged as the field journeys into a new era.

Quantifying in situ adaptive immune cell cognate interactions in humans.

Two-photon excitation microscopy (TPEM) has revolutionized the understanding of adaptive immunity. However, TPEM usually requires animal models and is not amenable to the study of human disease. The recognition of antigen by T cells requires cell contact and is associated with changes in T cell shape. We postulated that by capturing these features in fixed tissue samples, we could quantify in situ adaptive immunity. Therefore, we used a deep convolutional neural network to identify fundamental distance and cell-shape features associated with cognate help (cell-distance mapping (CDM)). In mice, CDM was comparable to TPEM in discriminating cognate T cell-dendritic cell (DC) interactions from non-cognate T cell-DC interactions. In human lupus nephritis, CDM confirmed that myeloid DCs present antigen to CD4+ T cells and identified plasmacytoid DCs as an important antigen-presenting cell. These data reveal a new approach with which to study human in situ adaptive immunity broadly applicable to autoimmunity, infection, and cancer.

Hyperactivated PI3Kδ promotes self and commensal reactivity at the expense of optimal humoral immunity.

Gain-of-function mutations in the gene encoding the phosphatidylinositol-3-OH kinase catalytic subunit p110δ (PI3Kδ) result in a human primary immunodeficiency characterized by lymphoproliferation, respiratory infections and inefficient responses to vaccines. However, what promotes these immunological disturbances at the cellular and molecular level remains unknown. We generated a mouse model that recapitulated major features of this disease and used this model and patient samples to probe how hyperactive PI3Kδ fosters aberrant humoral immunity. We found that mutant PI3Kδ led to co-stimulatory receptor ICOS-independent increases in the abundance of follicular helper T cells (TFH cells) and germinal-center (GC) B cells, disorganized GCs and poor class-switched antigen-specific responses to immunization, associated with altered regulation of the transcription factor FOXO1 and pro-apoptotic and anti-apoptotic members of the BCL-2 family. Notably, aberrant responses were accompanied by increased reactivity to gut bacteria and a broad increase in autoantibodies that were dependent on stimulation by commensal microbes. Our findings suggest that proper regulation of PI3Kδ is critical for ensuring optimal host-protective humoral immunity despite tonic stimulation from the commensal microbiome.

Healing the NIH-Funded Biomedical Research Enterprise.

Many feel that the R01 grant system supporting biomedical research in the U.S. is broken, discouraging entry of young investigators into the system and inadequately supporting more established investigators. Here, I argue for a "person-not-project"-based scheme that would permit creative, unfettered research by new investigators, better tie ongoing research contributions to continued funding, and help match the number of investigators seeking support with available funds.

Microbiota-Dependent Sequelae of Acute Infection Compromise Tissue-Specific Immunity.

Infections have been proposed as initiating factors for inflammatory disorders; however, identifying associations between defined infectious agents and the initiation of chronic disease has remained elusive. Here, we report that a single acute infection can have dramatic and long-term consequences for tissue-specific immunity. Following clearance of Yersinia pseudotuberculosis, sustained inflammation and associated lymphatic leakage in the mesenteric adipose tissue deviates migratory dendritic cells to the adipose compartment, thereby preventing their accumulation in the mesenteric lymph node. As a consequence, canonical mucosal immune functions, including tolerance and protective immunity, are persistently compromised. Post-resolution of infection, signals derived from the microbiota maintain inflammatory mesentery remodeling and consequently, transient ablation of the microbiota restores mucosal immunity. Our results indicate that persistent disruption of communication between tissues and the immune system following clearance of an acute infection represents an inflection point beyond which tissue homeostasis and immunity is compromised for the long-term. VIDEO ABSTRACT.

Niche-Specific Reprogramming of Epigenetic Landscapes Drives Myeloid Cell Diversity in Nonalcoholic Steatohepatitis.

Tissue-resident and recruited macrophages contribute to both host defense and pathology. Multiple macrophage phenotypes are represented in diseased tissues, but we lack deep understanding of mechanisms controlling diversification. Here, we investigate origins and epigenetic trajectories of hepatic macrophages during diet-induced non-alcoholic steatohepatitis (NASH). The NASH diet induced significant changes in Kupffer cell enhancers and gene expression, resulting in partial loss of Kupffer cell identity, induction of Trem2 and Cd9 expression, and cell death. Kupffer cell loss was compensated by gain of adjacent monocyte-derived macrophages that exhibited convergent epigenomes, transcriptomes, and functions. NASH-induced changes in Kupffer cell enhancers were driven by AP-1 and EGR that reprogrammed LXR functions required for Kupffer cell identity and survival to instead drive a scar-associated macrophage phenotype. These findings reveal mechanisms by which disease-associated environmental signals instruct resident and recruited macrophages to acquire distinct gene expression programs and corresponding functions.

Focusing in on T cell cross-reactivity.

To provide broad immunity to a vast array of foreign antigens with a limited number of T lymphocytes, each cell has to recognize many targets. By implementing a strategy to identify T cell receptor (TCR) ligands and investigating at a fine granularity their structure and sequence relationship, Birnbaum et al. demonstrate the surprisingly tight focus of such T cell cross-reactivity.

Global analyses of human immune variation reveal baseline predictors of postvaccination responses.

A major goal of systems biology is the development of models that accurately predict responses to perturbation. Constructing such models requires the collection of dense measurements of system states, yet transformation of data into predictive constructs remains a challenge. To begin to model human immunity, we analyzed immune parameters in depth both at baseline and in response to influenza vaccination. Peripheral blood mononuclear cell transcriptomes, serum titers, cell subpopulation frequencies, and B cell responses were assessed in 63 individuals before and after vaccination and were used to develop a systematic framework to dissect inter- and intra-individual variation and build predictive models of postvaccination antibody responses. Strikingly, independent of age and pre-existing antibody titers, accurate models could be constructed using pre-perturbation cell populations alone, which were validated using independent baseline time points. Most of the parameters contributing to prediction delineated temporally stable baseline differences across individuals, raising the prospect of immune monitoring before intervention.

The ß1-adrenergic receptor links sympathetic nerves to T cell exhaustion.

CD8+ T cells are essential components of the immune response against viral infections and tumours, and are capable of eliminating infected and cancerous cells. However, when the antigen cannot be cleared, T cells enter a state known as exhaustion1. Although it is clear that chronic antigen contributes to CD8+ T cell exhaustion, less is known about how stress responses in tissues regulate T cell function. Here we show a new link between the stress-associated catecholamines and the progression of T cell exhaustion through the ß1-adrenergic receptor ADRB1. We identify that exhausted CD8+ T cells increase ADRB1 expression and that exposure of ADRB1+ T cells to catecholamines suppresses their cytokine production and proliferation. Exhausted CD8+ T cells cluster around sympathetic nerves in an ADRB1-dependent manner. Ablation of ß1-adrenergic signalling limits the progression of T cells towards the exhausted state in chronic infection and improves effector functions when combined with immune checkpoint blockade (ICB) in melanoma. In a pancreatic cancer model resistant to ICB, ß-blockers and ICB synergize to boost CD8+ T cell responses and induce the development of tissue-resident memory-like T cells. Malignant disease is associated with increased catecholamine levels in patients2,3, and our results establish a connection between the sympathetic stress response, tissue innervation and T cell exhaustion. Here, we uncover a new mechanism by which blocking ß-adrenergic signalling in CD8+ T cells rejuvenates anti-tumour functions.

The Chemoattractant Receptor Ebi2 Drives Intranodal Naive CD4+ T Cell Peripheralization to Promote Effective Adaptive Immunity.

Lymph nodes (LNs) play critical roles in adaptive immunity by concentrating in one location the antigens, antigen-presenting cells, and antigen-responsive lymphocytes involved in such responses. Recent studies have revealed nonrandom localization of innate and adaptive immune cells within these organs, suggesting that microanatomical positioning optimizes responses involving sparse cooperating cells. Here, we report that the peripheral localization of LN cDC2 dendritic cells specialized for MHC-II antigen presentation is matched by a similarly biased paracortical distribution of CD4+ T cells directed by the chemoattractant receptor Ebi2. In the absence of Ebi2, CD4+ T cells lose their location bias and are delayed in antigen recognition, proliferative expansion, differentiation, direct effector activity, and provision of help for CD8+ T cell-mediated memory responses, limiting host defense and vaccine responses. These findings demonstrate evolutionary selection for distinct niches within the LN that promote cellular responses, emphasizing the critical link between fine-grained tissue organization and host defense.

A systems analysis identifies a feedforward inflammatory circuit leading to lethal influenza infection.

For acutely lethal influenza infections, the relative pathogenic contributions of direct viral damage to lung epithelium versus dysregulated immunity remain unresolved. Here, we take a top-down systems approach to this question. Multigene transcriptional signatures from infected lungs suggested that elevated activation of inflammatory signaling networks distinguished lethal from sublethal infections. Flow cytometry and gene expression analysis involving isolated cell subpopulations from infected lungs showed that neutrophil influx largely accounted for the predictive transcriptional signature. Automated imaging analysis, together with these gene expression and flow data, identified a chemokine-driven feedforward circuit involving proinflammatory neutrophils potently driven by poorly contained lethal viruses. Consistent with these data, attenuation, but not ablation, of the neutrophil-driven response increased survival without changing viral spread. These findings establish the primacy of damaging innate inflammation in at least some forms of influenza-induced lethality and provide a roadmap for the systematic dissection of infection-associated pathology.

The adaptor MAVS promotes NLRP3 mitochondrial localization and inflammasome activation.

NLRP3 is a key component of the macromolecular signaling complex called the inflammasome that promotes caspase 1-dependent production of IL-1ß. The adaptor ASC is necessary for NLRP3-dependent inflammasome function, but it is not known whether ASC is a sufficient partner and whether inflammasome formation occurs in the cytosol or in association with mitochondria is controversial. Here, we show that the mitochondria-associated adaptor molecule, MAVS, is required for optimal NLRP3 inflammasome activity. MAVS mediates recruitment of NLRP3 to mitochondria, promoting production of IL-1ß and the pathophysiologic activity of the NLRP3 inflammasome in vivo. Our data support a more complex model of NLRP3 inflammasome activation than previously appreciated, with at least two adapters required for maximal function. Because MAVS is a mitochondria-associated molecule previously considered to be uniquely involved in type 1 interferon production, these findings also reveal unexpected polygamous involvement of PYD/CARD-domain-containing adapters in innate immune signaling events.

Spatiotemporal basis of innate and adaptive immunity in secondary lymphoid tissue.

Secondary lymphoid tissues are the sites of both innate and adaptive host defense. Aside from the relatively static nonhematopoietic stromal elements and some macrophages and dendritic cells, most of the cells in these tissues are in constant movement, but the organs maintain a defined microanatomy with preferred locations for the bulk of T cells, B cells, and other lymphocytes and subsets of myeloid cells. Here we describe both the cell dynamics and spatial organization of lymph nodes and review how both physical features and molecular cues guide cell movement to optimize host defense. We emphasize the role of locality in improving the efficiency of a system requiring rare cells to find each other and interact productively through membrane-bound or short-range secreted mediators and highlight how changes in steady-state cell positioning during an infectious challenge contribute to rapid generation of productive responses.

A spatially-organized multicellular innate immune response in lymph nodes limits systemic pathogen spread.

The lymphatic network that transports interstitial fluid and antigens to lymph nodes constitutes a conduit system that can be hijacked by invading pathogens to achieve systemic spread unless dissemination is blocked in the lymph node itself. Here, we show that a network of diverse lymphoid cells (natural killer cells, γδ T cells, natural killer T cells, and innate-like CD8+ T cells) are spatially prepositioned close to lymphatic sinus-lining sentinel macrophages where they can rapidly and efficiently receive inflammasome-generated IL-18 and additional cytokine signals from the pathogen-sensing phagocytes. This leads to rapid IFNγ secretion by the strategically positioned innate lymphocytes, fostering antimicrobial resistance in the macrophage population. Interference with this innate immune response loop allows systemic spread of lymph-borne bacteria. These findings extend our understanding of the functional significance of cellular positioning and local intercellular communication within lymph nodes while emphasizing the role of these organs as highly active locations of innate host defense.

Author Correction: Commensal-driven immune zonation of the liver promotes host defence.

A Correction to this paper has been published: https://doi.org/10.1038/s41586-021-03346-0.

Commensal-driven immune zonation of the liver promotes host defence.

The liver connects the intestinal portal vasculature with the general circulation, using a diverse array of immune cells to protect from pathogens that translocate from the gut1. In liver lobules, blood flows from portal triads that are situated in periportal lobular regions to the central vein via a polarized sinusoidal network. Despite this asymmetry, resident immune cells in the liver are considered to be broadly dispersed across the lobule. This differs from lymphoid organs, in which immune cells adopt spatially biased positions to promote effective host defence2,3. Here we used quantitative multiplex imaging, genetic perturbations, transcriptomics, infection-based assays and mathematical modelling to reassess the relationship between the localization of immune cells in the liver and host protection. We found that myeloid and lymphoid resident immune cells concentrate around periportal regions. This asymmetric localization was not developmentally controlled, but resulted from sustained MYD88-dependent signalling induced by commensal bacteria in liver sinusoidal endothelial cells, which in turn regulated the composition of the pericellular matrix involved in the formation of chemokine gradients. In vivo experiments and modelling showed that this immune spatial polarization was more efficient than a uniform distribution in protecting against systemic bacterial dissemination. Together, these data reveal that liver sinusoidal endothelial cells sense the microbiome, actively orchestrating the localization of immune cells, to optimize host defence.