A unified metric of human immune health.

Immunological health has been challenging to characterize but could be defined as the absence of immune pathology. While shared features of some immune diseases and the concept of immunologic resilience based on age-independent adaptation to antigenic stimulation have been developed, general metrics of immune health and its utility for assessing clinically healthy individuals remain ill defined. Here we integrated transcriptomics, serum protein, peripheral immune cell frequency and clinical data from 228 patients with 22 monogenic conditions impacting key immunological pathways together with 42 age- and sex-matched healthy controls. Despite the high penetrance of monogenic lesions, differences between individuals in diverse immune parameters tended to dominate over those attributable to disease conditions or medication use. Unsupervised or supervised machine learning independently identified a score that distinguished healthy participants from patients with monogenic diseases, thus suggesting a quantitative immune health metric (IHM). In ten independent datasets, the IHM discriminated healthy from polygenic autoimmune and inflammatory disease states, marked aging in clinically healthy individuals, tracked disease activities and treatment responses in both immunological and nonimmunological diseases, and predicted age-dependent antibody responses to immunizations with different vaccines. This discriminatory power goes beyond that of the classical inflammatory biomarkers C-reactive protein and interleukin-6. Thus, deviations from health in diverse conditions, including aging, have shared systemic immune consequences, and we provide a web platform for calculating the IHM for other datasets, which could empower precision medicine.

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.

Preliminary findings from the Dynamics of the Immune Responses to Repeat Influenza Vaccination Exposures (DRIVE I) Study: a Randomized Controlled Trial.

Background: Studies have reported that repeated annual vaccination may influence the effectiveness of the influenza vaccination in the current season. The mechanisms underlying these differences are unclear but might include "focusing" of the adaptive immune response to older strains. Methods: We established a 5-year randomized placebo-controlled trial of repeated influenza vaccination (Flublok, Sanofi Pasteur) in adults 18-45 years of age. Participants were randomized equally between five groups, with planned annual receipt of vaccination (V) or saline placebo (P) as follows: P-P-P-P-V, P-P-P-V-V, P-P-V-V-V, P-V-V-V-V, or V-V-V-VV. Serum samples were collected each year just before vaccination and after 30 and 182 days. A subset of sera were tested by hemagglutination inhibition assays, focus reduction neutralization tests and enzyme-linked immunosorbent assays against vaccine strains. Results: From 23 October 2020 through 11 March 2021 we enrolled and randomized 447 adults. We selected sera from 95 participants at five timepoints from the first two study years for testing. Among vaccinated individuals, antibody titers increased between days 0 and 30 against each of the vaccine strains, with substantial increases for first-time vaccinees and smaller increases for repeat vaccinees, who had higher pre-vaccination titers in year 2. There were statistically significant reductions in the proportion of participants achieving a four-fold greater rise in antibody titer for the repeat vaccinees for A(H1N1), B/Victoria and B/Yamagata, but not for influenza A(H3N2). There were no statistically significant differences between groups in geometric mean titers at day 30 or the proportions of participants with antibody titers ≥40 at day 30 for any of the vaccine strains. Conclusions: In the first two years, repeat vaccinees and first-time vaccinees had similar post-vaccination geometric mean titers to all four vaccine strains, indicative of similar levels of clinical protection. The vaccine strains of A(H1N1) and A(H3N2) were updated in year 2, providing an opportunity to explore antigenic distances between those strains in humans in subsequent years.

Integrating population and single-cell variations in vaccine responses identifies a naturally adjuvanted human immune setpoint.

Multimodal single-cell profiling methods can capture immune cell variations unfolding over time at the molecular, cellular, and population levels. Transforming these data into biological insights remains challenging. Here, we introduce a framework to integrate variations at the human population and single-cell levels in vaccination responses. Comparing responses following AS03-adjuvanted versus unadjuvanted influenza vaccines with CITE-seq revealed AS03-specific early (day 1) response phenotypes, including a B cell signature of elevated germinal center competition. A correlated network of cell-type-specific transcriptional states defined the baseline immune status associated with high antibody responders to the unadjuvanted vaccine. Certain innate subsets in the network appeared "naturally adjuvanted," with transcriptional states resembling those induced uniquely by AS03-adjuvanted vaccination. Consistently, CD14+ monocytes from high responders at baseline had elevated phospho-signaling responses to lipopolysaccharide stimulation. Our findings link baseline immune setpoints to early vaccine responses, with positive implications for adjuvant development and immune response engineering.

Characterization of the antispike IgG immune response to COVID-19 vaccines in people with a wide variety of immunodeficiencies.

Research on coronavirus disease 2019 vaccination in immune-deficient/disordered people (IDP) has focused on cancer and organ transplantation populations. In a prospective cohort of 195 IDP and 35 healthy volunteers (HV), antispike immunoglobulin G (IgG) was detected in 88% of IDP after dose 2, increasing to 93% by 6 months after dose 3. Despite high seroconversion, median IgG levels for IDP never surpassed one-third that of HV. IgG binding to Omicron BA.1 was lowest among variants. Angiotensin-converting enzyme 2 pseudo-neutralization only modestly correlated with antispike IgG concentration. IgG levels were not significantly altered by receipt of different messenger RNA-based vaccines, immunomodulating treatments, and prior severe acute respiratory syndrome coronavirus 2 infections. While our data show that three doses of coronavirus disease 2019 vaccinations induce antispike IgG in most IDP, additional doses are needed to increase protection. Because of the notably reduced IgG response to Omicron BA.1, the efficacy of additional vaccinations, including bivalent vaccines, should be studied in this population.

Systems immunology of regulatory T cells: can one circuit explain it all?

Regulatory T (Treg) cells play vital roles in immune homeostasis and response, including discrimination between self- and non-self-antigens, containment of immunopathology, and inflammation resolution. These diverse functions are orchestrated by cellular circuits involving Tregs and other cell types across space and time. Despite dramatic progress in our understanding of Treg biology, a quantitative framework capturing how Treg-containing circuits give rise to these diverse functions is lacking. Here, we propose that different facets of Treg function can be interpreted as distinct operating regimes of the same underlying circuit. We discuss how a systems immunology approach, involving quantitative experiments, computational modeling, and machine learning, can advance our understanding of Treg function, and help identify general operating and design principles underlying immune regulation.

Non-coding sequence variation reveals fragility within interleukin 2 feedback circuitry and shapes autoimmune disease risk.

Genetic variants associated with human autoimmune diseases commonly map to non-coding control regions, particularly enhancers that function selectively in immune cells and fine-tune gene expression within a relatively narrow range of values. How such modest, cell-type-selective changes can meaningfully shape organismal disease risk remains unclear. To explore this issue, we experimentally manipulated species-conserved enhancers within the disease-associated IL2RA locus and studied accompanying changes in the progression of autoimmunity. Perturbing distinct enhancers with restricted activity in conventional T cells (Tconvs) or regulatory T cells (Tregs)-two functionally antagonistic T cell subsets-caused only modest, cell-type-selective decreases in IL2ra expression parameters. However, these same perturbations had striking and opposing effects in vivo , completely preventing or severely accelerating disease in a murine model of type 1 diabetes. Quantitative tissue imaging and computational modelling revealed that each enhancer manipulation impinged on distinct IL-2-dependent feedback circuits. These imbalances altered the intracellular signaling and intercellular communication dynamics of activated Tregs and Tconvs, producing opposing spatial domains that amplified or constrained ongoing autoimmune responses. These findings demonstrate how subtle changes in gene regulation stemming from non-coding variation can propagate across biological scales due to non-linearities in intra- and intercellular feedback circuitry, dramatically shaping disease risk at the organismal level.

Tracking B cell responses to the SARS-CoV-2 mRNA-1273 vaccine.

Protective immunity following vaccination is sustained by long-lived antibody-secreting cells and resting memory B cells (MBCs). Responses to two-dose SARS-CoV-2 mRNA-1273 vaccination are evaluated longitudinally by multimodal single-cell analysis in three infection-naïve individuals. Integrated surface protein, transcriptomics, and B cell receptor (BCR) repertoire analysis of sorted plasmablasts and spike+ (S-2P+) and S-2P- B cells reveal clonal expansion and accumulating mutations among S-2P+ cells. These cells are enriched in a cluster of immunoglobulin G-expressing MBCs and evolve along a bifurcated trajectory rooted in CXCR3+ MBCs. One branch leads to CD11c+ atypical MBCs while the other develops from CD71+ activated precursors to resting MBCs, the dominant population at month 6. Among 12 evolving S-2P+ clones, several are populated with plasmablasts at early timepoints as well as CD71+ activated and resting MBCs at later timepoints, and display intra- and/or inter-cohort BCR convergence. These relationships suggest a coordinated and predictable evolution of SARS-CoV-2 vaccine-generated MBCs.

Inflammatory and interferon gene expression signatures in patients with mitochondrial disease.

BACKGROUND: People with mitochondrial disease (MtD) are susceptible to metabolic decompensation and neurological symptom progression in response to an infection. Increasing evidence suggests that mitochondrial dysfunction may cause chronic inflammation, which may promote hyper-responsiveness to pathogens and neurodegeneration. We sought to examine transcriptional changes between MtD patients and healthy controls to identify common gene signatures of immune dysregulation in MtD. METHODS: We collected whole blood from a cohort of MtD patients and healthy controls and performed RNAseq to examine transcriptomic differences. We performed GSEA analyses to compare our findings against existing studies to identify commonly dysregulated pathways. RESULTS: Gene sets involved in inflammatory signaling, including type I interferons, interleukin-1ß and antiviral responses, are enriched in MtD patients compared to controls. Monocyte and dendritic cell gene clusters are also enriched in MtD patients, while T cell and B cell gene sets are negatively enriched. The enrichment of antiviral response corresponds with an independent set of MELAS patients, and two mouse models of mtDNA dysfunction. CONCLUSIONS: Through the convergence of our results, we demonstrate translational evidence of systemic peripheral inflammation arising from MtD, predominantly through antiviral response gene sets. This provides key evidence linking mitochondrial dysfunction to inflammation, which may contribute to the pathogenesis of primary MtD and other chronic inflammatory disorders associated with mitochondrial dysfunction.

Multiscale integration of human and single-cell variations reveals unadjuvanted vaccine high responders are naturally adjuvanted.

Advances in multimodal single cell analysis can empower high-resolution dissection of human vaccination responses. The resulting data capture multiple layers of biological variations, including molecular and cellular states, vaccine formulations, inter- and intra-subject differences, and responses unfolding over time. Transforming such data into biological insight remains a major challenge. Here we present a systematic framework applied to multimodal single cell data obtained before and after influenza vaccination without adjuvants or pandemic H5N1 vaccination with the AS03 adjuvant. Our approach pinpoints responses shared across or unique to specific cell types and identifies adjuvant specific signatures, including pro-survival transcriptional states in B lymphocytes that emerged one day after vaccination. We also reveal that high antibody responders to the unadjuvanted vaccine have a distinct baseline involving a rewired network of cell type specific transcriptional states. Remarkably, the status of certain innate immune cells in this network in high responders of the unadjuvanted vaccine appear "naturally adjuvanted": they resemble phenotypes induced early in the same cells only by vaccination with AS03. Furthermore, these cell subsets have elevated frequency in the blood at baseline and increased cell-intrinsic phospho-signaling responses after LPS stimulation ex vivo in high compared to low responders. Our findings identify how variation in the status of multiple immune cell types at baseline may drive robust differences in innate and adaptive responses to vaccination and thus open new avenues for vaccine development and immune response engineering in humans.

Inflammatory and interferon gene expression signatures in patients with mitochondrial disease.

Background: People with mitochondrial disease (MtD) are susceptible to metabolic decompensation and neurological symptom progression in response to an infection. Increasing evidence suggests that mitochondrial dysfunction may cause chronic inflammation, which may promote hyperresponsiveness to pathogens and neurodegeneration. Methods: We collected whole blood from a cohort of MtD patients and healthy controls and performed RNAseq to examine transcriptomic differences. We performed GSEA analyses to compare our findings against existing studies to identify commonly dysregulated pathways. Results: Gene sets involved in inflammatory signaling, including type I interferons, interleukin-1ß and antiviral responses, are enriched in MtD patients compared to controls. Monocyte and dendritic cell gene clusters are also enriched in MtD patients, while T cell and B cell gene sets are negatively enriched. The enrichment of antiviral response corresponds with an independent set of MELAS patients, and two mouse models of mtDNA dysfunction. Conclusions: Through the convergence of our results, we demonstrate translational evidence of systemic peripheral inflammation arising from MtD, predominantly through antiviral response gene sets. This provides key evidence linking mitochondrial dysfunction to inflammation, which may contribute to the pathogenesis of primary MtD and other chronic inflammatory disorders associated with mitochondrial dysfunction.

Multiomics integration of 22 immune-mediated monogenic diseases reveals an emergent axis of human immune health.

Monogenic diseases are often studied in isolation due to their rarity. Here we utilize multiomics to assess 22 monogenic immune-mediated conditions with age- and sex-matched healthy controls. Despite clearly detectable disease-specific and "pan-disease" signatures, individuals possess stable personal immune states over time. Temporally stable differences among subjects tend to dominate over differences attributable to disease conditions or medication use. Unsupervised principal variation analysis of personal immune states and machine learning classification distinguishing between healthy controls and patients converge to a metric of immune health (IHM). The IHM discriminates healthy from multiple polygenic autoimmune and inflammatory disease states in independent cohorts, marks healthy aging, and is a pre-vaccination predictor of antibody responses to influenza vaccination in the elderly. We identified easy-to-measure circulating protein biomarker surrogates of the IHM that capture immune health variations beyond age. Our work provides a conceptual framework and biomarkers for defining and measuring human immune health.

Sex and prior exposure jointly shape innate immune responses to a live herpesvirus vaccine.

Background: Both sex and prior exposure to pathogens are known to influence responses to immune challenges, but their combined effects are not well established in humans, particularly in early innate responses critical for shaping subsequent outcomes. Methods: We employed systems immunology approaches to study responses to a replication-defective, herpes simplex virus (HSV) 2 vaccine in men and women either naive or previously exposed to HSV. Results: Blood transcriptomic and cell population profiling showed substantial changes on day 1 after vaccination, but the responses depended on sex and whether the vaccinee was naive or previously exposed to HSV. The magnitude of early transcriptional responses was greatest in HSV naive women where type I interferon (IFN) signatures were prominent and associated negatively with vaccine-induced neutralizing antibody titers, suggesting that a strong early antiviral response reduced the uptake of this replication-defective virus vaccine. While HSV seronegative vaccine recipients had upregulation of gene sets in type I IFN (IFN-α/ß) responses, HSV2 seropositive vaccine recipients tended to have responses focused more on type II IFN (IFN-γ) genes. Conclusions: These results together show that prior exposure and sex interact to shape early innate responses that then impact subsequent adaptive immune phenotypes. Funding: Intramural Research Program of the NIH, the National Institute of Allergy and Infectious Diseases, and other institutes supporting the Trans-NIH Center for Human Immunology, Autoimmunity, and Inflammation. The vaccine trial was supported through a clinical trial agreement between the National Institute of Allergy and Infectious Diseases and Sanofi Pasteur. Clinical trial number: NCT01915212.

Influenza vaccination reveals sex dimorphic imprints of prior mild COVID-19.

Acute viral infections can have durable functional impacts on the immune system long after recovery, but how they affect homeostatic immune states and responses to future perturbations remain poorly understood1-4. Here we use systems immunology approaches, including longitudinal multimodal single-cell analysis (surface proteins, transcriptome and V(D)J sequences) to comparatively assess baseline immune statuses and responses to influenza vaccination in 33 healthy individuals after recovery from mild, non-hospitalized COVID-19 (mean, 151 days after diagnosis) and 40 age- and sex-matched control individuals who had never had COVID-19. At the baseline and independent of time after COVID-19, recoverees had elevated T cell activation signatures and lower expression of innate immune genes including Toll-like receptors in monocytes. Male individuals who had recovered from COVID-19 had coordinately higher innate, influenza-specific plasmablast, and antibody responses after vaccination compared with healthy male individuals and female individuals who had recovered from COVID-19, in part because male recoverees had monocytes with higher IL-15 responses early after vaccination coupled with elevated prevaccination frequencies of 'virtual memory'-like CD8+ T cells poised to produce more IFNγ after IL-15 stimulation. Moreover, the expression of the repressed innate immune genes in monocytes increased by day 1 to day 28 after vaccination in recoverees, therefore moving towards the prevaccination baseline of the healthy control individuals. By contrast, these genes decreased on day 1 and returned to the baseline by day 28 in the control individuals. Our study reveals sex-dimorphic effects of previous mild COVID-19 and suggests that viral infections in humans can establish new immunological set-points that affect future immune responses in an antigen-agnostic manner.

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.

Transcriptional atlas of the human immune response to 13 vaccines reveals a common predictor of vaccine-induced antibody responses.

Systems vaccinology has defined molecular signatures and mechanisms of immunity to vaccination. However, comparative analysis of immunity to different vaccines is lacking. We integrated transcriptional data of over 3,000 samples, from 820 adults across 28 studies of 13 vaccines and analyzed vaccination-induced signatures of antibody responses. Most vaccines induced signatures of innate immunity and plasmablasts at days 1 and 7, respectively, after vaccination. However, the yellow fever vaccine induced an early transient signature of T and B cell activation at day 1, followed by delayed antiviral/interferon and plasmablast signatures that peaked at days 7 and 14-21, respectively. Thus, there was no evidence for a 'universal signature' that predicted antibody response to all vaccines. However, accounting for the asynchronous nature of responses, we defined a time-adjusted signature that predicted antibody responses across vaccines. These results provide a transcriptional atlas of immunity to vaccination and define a common, time-adjusted signature of antibody responses.

Pan-vaccine analysis reveals innate immune endotypes predictive of antibody responses to vaccination.

Several studies have shown that the pre-vaccination immune state is associated with the antibody response to vaccination. However, the generalizability and mechanisms that underlie this association remain poorly defined. Here, we sought to identify a common pre-vaccination signature and mechanisms that could predict the immune response across 13 different vaccines. Analysis of blood transcriptional profiles across studies revealed three distinct pre-vaccination endotypes, characterized by the differential expression of genes associated with a pro-inflammatory response, cell proliferation, and metabolism alterations. Importantly, individuals whose pre-vaccination endotype was enriched in pro-inflammatory response genes known to be downstream of nuclear factor-kappa B showed significantly higher serum antibody responses 1 month after vaccination. This pro-inflammatory pre-vaccination endotype showed gene expression characteristic of the innate activation state triggered by Toll-like receptor ligands or adjuvants. These results demonstrate that wide variations in the transcriptional state of the immune system in humans can be a key determinant of responsiveness to vaccination.

The Immune Signatures data resource, a compendium of systems vaccinology datasets.

Vaccines are among the most cost-effective public health interventions for preventing infection-induced morbidity and mortality, yet much remains to be learned regarding the mechanisms by which vaccines protect. Systems immunology combines traditional immunology with modern 'omic profiling techniques and computational modeling to promote rapid and transformative advances in vaccinology and vaccine discovery. The NIH/NIAID Human Immunology Project Consortium (HIPC) has leveraged systems immunology approaches to identify molecular signatures associated with the immunogenicity of many vaccines. However, comparative analyses have been limited by the distributed nature of some data, potential batch effects across studies, and the absence of multiple relevant studies from non-HIPC groups in ImmPort. To support comparative analyses across different vaccines, we have created the Immune Signatures Data Resource, a compendium of standardized systems vaccinology datasets. This data resource is available through ImmuneSpace, along with code to reproduce the processing and batch normalization starting from the underlying study data in ImmPort and the Gene Expression Omnibus (GEO). The current release comprises 1405 participants from 53 cohorts profiling the response to 24 different vaccines. This novel systems vaccinology data release represents a valuable resource for comparative and meta-analyses that will accelerate our understanding of mechanisms underlying vaccine responses.

Peroxisome proliferator activated receptor-γ agonist pioglitazone improves vascular and metabolic dysfunction in systemic lupus erythematosus.

OBJECTIVES: Premature cardiovascular events in systemic lupus erythematosus (SLE) contribute to morbidity and mortality, with no effective preventive strategies described to date. Immune dysregulation and metabolic disturbances appear to play prominent roles in the induction of vascular disease in SLE. The peroxisome proliferator activated receptor-gamma agonist pioglitazone (PGZ suppresses vascular damage and immune dysregulation in murine lupus and improves endothelial dysfunction in other inflammatory diseases. We hypothesised that PGZ could improve vascular dysfunction and cardiometabolic parameters in SLE. METHODS: Eighty SLE subjects with mild to severe disease activity were randomised to a sequence of PGZ followed by placebo for 3 months, or vice versa, in a double-blind, cross-over design with a 2-month wash-out period. Primary endpoints were parameters of endothelial function and arterial inflammation, measured by multimodal assessments. Additional outcome measures of disease activity, neutrophil dysregulation, metabolic disturbances and gene expression studies were performed. RESULTS: Seventy-two subjects completed the study. PGZ was associated with a significant reduction in Cardio-Ankle Vascular Index (a measure of arterial stiffness) compared with placebo. Various metabolic parameters improved with PGZ, including insulin resistance and lipoprotein profiles. Circulating neutrophil extracellular trap levels also significantly decreased with PGZ compared with placebo. Most adverse events experienced while on PGZ were mild and resolved with reduction in PGZ dose. CONCLUSION: PGZ was well tolerated and induced significant improvement in vascular stiffness and cardiometabolic parameters in SLE. The results suggest that PGZ should be further explored as a modulator of cardiovascular disease risk in SLE. TRIAL REGISTRATION NUMBER: NCT02338999.

Considering innate immune responses in SARS-CoV-2 infection and COVID-19.

During the COVID-19 pandemic, much of the media focus has been on adaptive immunity, particularly antibody levels and memory T cells. However, immunologists have been striving to decipher how SARS-CoV-2 infection impacts our first line of defence, namely the innate immune system. In early 2022, Program staff from the NIAID at the NIH organized a workshop focusing on the innate immune response to SARS-CoV-2 infection and during COVID-19, which was chaired by Ralph Baric, Jenny Ting and John Lambris. Following the meeting, Nature Reviews Immunology invited some of the organizers and speakers to share their thoughts on the key discussion points.