Deconstruction of rheumatoid arthritis synovium defines inflammatory subtypes.

Rheumatoid arthritis is a prototypical autoimmune disease that causes joint inflammation and destruction1. There is currently no cure for rheumatoid arthritis, and the effectiveness of treatments varies across patients, suggesting an undefined pathogenic diversity1,2. Here, to deconstruct the cell states and pathways that characterize this pathogenic heterogeneity, we profiled the full spectrum of cells in inflamed synovium from patients with rheumatoid arthritis. We used multi-modal single-cell RNA-sequencing and surface protein data coupled with histology of synovial tissue from 79 donors to build single-cell atlas of rheumatoid arthritis synovial tissue that includes more than 314,000 cells. We stratified tissues into six groups, referred to as cell-type abundance phenotypes (CTAPs), each characterized by selectively enriched cell states. These CTAPs demonstrate the diversity of synovial inflammation in rheumatoid arthritis, ranging from samples enriched for T and B cells to those largely lacking lymphocytes. Disease-relevant cell states, cytokines, risk genes, histology and serology metrics are associated with particular CTAPs. CTAPs are dynamic and can predict treatment response, highlighting the clinical utility of classifying rheumatoid arthritis synovial phenotypes. This comprehensive atlas and molecular, tissue-based stratification of rheumatoid arthritis synovial tissue reveal new insights into rheumatoid arthritis pathology and heterogeneity that could inform novel targeted treatments.

Immunosuppression causes dynamic changes in expression QTLs in psoriatic skin.

Psoriasis is a chronic, systemic inflammatory condition primarily affecting skin. While the role of the immune compartment (e.g., T cells) is well established, the changes in the skin compartment are more poorly understood. Using longitudinal skin biopsies (n = 375) from the "Psoriasis Treatment with Abatacept and Ustekinumab: A Study of Efficacy"(PAUSE) clinical trial (n = 101), we report 953 expression quantitative trait loci (eQTLs). Of those, 116 eQTLs have effect sizes that were modulated by local skin inflammation (eQTL interactions). By examining these eQTL genes (eGenes), we find that most are expressed in the skin tissue compartment, and a subset overlap with the NRF2 pathway. Indeed, the strongest eQTL interaction signal - rs1491377616-LCE3C - links a psoriasis risk locus with a gene specifically expressed in the epidermis. This eQTL study highlights the potential to use biospecimens from clinical trials to discover in vivo eQTL interactions with therapeutically relevant environmental variables.

Granzyme K+ CD8 T cells form a core population in inflamed human tissue.

T cell-derived pro-inflammatory cytokines are a major driver of rheumatoid arthritis (RA) pathogenesis. Although these cytokines have traditionally been attributed to CD4 T cells, we have found that CD8 T cells are notably abundant in synovium and make more interferon (IFN)-γ and nearly as much tumor necrosis factor (TNF) as their CD4 T cell counterparts. Furthermore, using unbiased high-dimensional single-cell RNA-seq and flow cytometric data, we found that the vast majority of synovial tissue and synovial fluid CD8 T cells belong to an effector CD8 T cell population characterized by high expression of granzyme K (GzmK) and low expression of granzyme B (GzmB) and perforin. Functional experiments demonstrate that these GzmK+ GzmB+ CD8 T cells are major cytokine producers with low cytotoxic potential. Using T cell receptor repertoire data, we found that CD8 GzmK+ GzmB+ T cells are clonally expanded in synovial tissues and maintain their granzyme expression and overall cell state in blood, suggesting that they are enriched in tissue but also circulate. Using GzmK and GzmB signatures, we found that GzmK-expressing CD8 T cells were also the major CD8 T cell population in the gut, kidney, and coronavirus disease 2019 (COVID-19) bronchoalveolar lavage fluid, suggesting that they form a core population of tissue-associated T cells across diseases and human tissues. We term this population tissue-enriched expressing GzmK or TteK CD8 cells. Armed to produce cytokines in response to both antigen-dependent and antigen-independent stimuli, CD8 TteK cells have the potential to drive inflammation.

FcγR engagement reprograms neutrophils into antigen cross-presenting cells that elicit acquired anti-tumor immunity.

Classical dendritic cells (cDC) are professional antigen-presenting cells (APC) that regulate immunity and tolerance. Neutrophil-derived cells with properties of DCs (nAPC) are observed in human diseases and after culture of neutrophils with cytokines. Here we show that FcγR-mediated endocytosis of antibody-antigen complexes or an anti-FcγRIIIB-antigen conjugate converts neutrophils into nAPCs that, in contrast to those generated with cytokines alone, activate T cells to levels observed with cDCs and elicit CD8+ T cell-dependent anti-tumor immunity in mice. Single cell transcript analyses and validation studies implicate the transcription factor PU.1 in neutrophil to nAPC conversion. In humans, blood nAPC frequency in lupus patients correlates with disease. Moreover, anti-FcγRIIIB-antigen conjugate treatment induces nAPCs that can activate autologous T cells when using neutrophils from individuals with myeloid neoplasms that harbor neoantigens or those vaccinated against bacterial toxins. Thus, anti-FcγRIIIB-antigen conjugate-induced conversion of neutrophils to immunogenic nAPCs may represent a possible immunotherapy for cancer and infectious diseases.

IFN-γ and TNF-α drive a CXCL10+ CCL2+ macrophage phenotype expanded in severe COVID-19 lungs and inflammatory diseases with tissue inflammation.

BACKGROUND: Immunosuppressive and anti-cytokine treatment may have a protective effect for patients with COVID-19. Understanding the immune cell states shared between COVID-19 and other inflammatory diseases with established therapies may help nominate immunomodulatory therapies. METHODS: To identify cellular phenotypes that may be shared across tissues affected by disparate inflammatory diseases, we developed a meta-analysis and integration pipeline that models and removes the effects of technology, tissue of origin, and donor that confound cell-type identification. Using this approach, we integrated > 300,000 single-cell transcriptomic profiles from COVID-19-affected lungs and tissues from healthy subjects and patients with five inflammatory diseases: rheumatoid arthritis (RA), Crohn's disease (CD), ulcerative colitis (UC), systemic lupus erythematosus (SLE), and interstitial lung disease. We tested the association of shared immune states with severe/inflamed status compared to healthy control using mixed-effects modeling. To define environmental factors within these tissues that shape shared macrophage phenotypes, we stimulated human blood-derived macrophages with defined combinations of inflammatory factors, emphasizing in particular antiviral interferons IFN-beta (IFN-ß) and IFN-gamma (IFN-γ), and pro-inflammatory cytokines such as TNF. RESULTS: We built an immune cell reference consisting of > 300,000 single-cell profiles from 125 healthy or disease-affected donors from COVID-19 and five inflammatory diseases. We observed a CXCL10+ CCL2+ inflammatory macrophage state that is shared and strikingly abundant in severe COVID-19 bronchoalveolar lavage samples, inflamed RA synovium, inflamed CD ileum, and UC colon. These cells exhibited a distinct arrangement of pro-inflammatory and interferon response genes, including elevated levels of CXCL10, CXCL9, CCL2, CCL3, GBP1, STAT1, and IL1B. Further, we found this macrophage phenotype is induced upon co-stimulation by IFN-γ and TNF-α. CONCLUSIONS: Our integrative analysis identified immune cell states shared across inflamed tissues affected by inflammatory diseases and COVID-19. Our study supports a key role for IFN-γ together with TNF-α in driving an abundant inflammatory macrophage phenotype in severe COVID-19-affected lungs, as well as inflamed RA synovium, CD ileum, and UC colon, which may be targeted by existing immunomodulatory therapies.

IFN- γ and TNF- α drive a CXCL10 + CCL2 + macrophage phenotype expanded in severe COVID-19 and other diseases with tissue inflammation.

Immunosuppressive and anti-cytokine treatment may have a protective effect for patients with COVID-19. Understanding the immune cell states shared between COVID-19 and other inflammatory diseases with established therapies may help nominate immunomodulatory therapies. Using an integrative strategy, we built a reference by meta-analyzing > 300,000 immune cells from COVID-19 and 5 inflammatory diseases including rheumatoid arthritis (RA), Crohn's disease (CD), ulcerative colitis (UC), lupus, and interstitial lung disease. Our cross-disease analysis revealed that an FCN1 + inflammatory macrophage state is common to COVID-19 bronchoalveolar lavage samples, RA synovium, CD ileum, and UC colon. We also observed that a CXCL10 + CCL2 + inflammatory macrophage state is abundant in severe COVID-19, inflamed CD and RA, and expresses inflammatory genes such as GBP1, STAT1 , and IL1B . We found that the CXCL10 + CCL2 + macrophages are transcriptionally similar to blood-derived macrophages stimulated with TNF- α and IFN- γ ex vivo . Our findings suggest that IFN- γ , alongside TNF- α , might be a key driver of this abundant inflammatory macrophage phenotype in severe COVID-19 and other inflammatory diseases, which may be targeted by existing immunomodulatory therapies.