Immune checkpoint inhibitor-induced colitis is mediated by polyfunctional lymphocytes and is dependent on an IL23/IFNγ axis.

Immune checkpoint inhibitors (CPIs) are a relatively newly licenced cancer treatment, which make a once previously untreatable disease now amenable to a potential cure. Combination regimens of anti-CTLA4 and anti-PD-1 show enhanced efficacy but are prone to off-target immune-mediated tissue injury, particularly at the barrier surfaces. To probe the impact of immune checkpoints on intestinal homoeostasis, mice are challenged with anti-CTLA4 and anti-PD-1 immunotherapy and manipulation of the intestinal microbiota. The immune profile of the colon of these mice with CPI-colitis is analysed using bulk RNA sequencing, single-cell RNA sequencing and flow cytometry. CPI-colitis in mice is dependent on the composition of the intestinal microbiota and by the induction of lymphocytes expressing interferon-γ (IFNγ), cytotoxicity molecules and other pro-inflammatory cytokines/chemokines. This pre-clinical model of CPI-colitis could be attenuated following blockade of the IL23/IFNγ axis. Therapeutic targeting of IFNγ-producing lymphocytes or regulatory networks, may hold the key to reversing CPI-colitis.

CD90 is not constitutively expressed in functional innate lymphoid cells.

Huge progress has been made in understanding the biology of innate lymphoid cells (ILC) by adopting several well-known concepts in T cell biology. As such, flow cytometry gating strategies and markers, such as CD90, have been applied to indentify ILC. Here, we report that most non-NK intestinal ILC have a high expression of CD90 as expected, but surprisingly a sub-population of cells exhibit low or even no expression of this marker. CD90-negative and CD90-low CD127+ ILC were present amongst all ILC subsets in the gut. The frequency of CD90-negative and CD90-low CD127+ ILC was dependent on stimulatory cues in vitro and enhanced by dysbiosis in vivo. CD90-negative and CD90-low CD127+ ILC were a potential source of IL-13, IFNγ and IL-17A at steady state and upon dysbiosis- and dextran sulphate sodium-elicited colitis. Hence, this study reveals that, contrary to expectations, CD90 is not constitutively expressed by functional ILC in the gut.

Monocyte migration profiles define disease severity in acute COVID-19 and unique features of long COVID.

BACKGROUND: COVID-19 is associated with a dysregulated immune response but it is unclear how immune dysfunction contributes to the chronic morbidity persisting in many COVID-19 patients during convalescence (long COVID). METHODS: We assessed phenotypical and functional changes of monocytes in COVID-19 patients during hospitalisation and up to 9 months of convalescence following COVID-19, respiratory syncytial virus or influenza A. Patients with progressive fibrosing interstitial lung disease were included as a positive control for severe, ongoing lung injury. RESULTS: Monocyte alterations in acute COVID-19 patients included aberrant expression of leukocyte migration molecules, continuing into convalescence (n=142) and corresponding with specific symptoms of long COVID. Long COVID patients with unresolved lung injury, indicated by sustained shortness of breath and abnormal chest radiology, were defined by high monocyte expression of C-X-C motif chemokine receptor 6 (CXCR6) (p<0.0001) and adhesion molecule P-selectin glycoprotein ligand 1 (p<0.01), alongside preferential migration of monocytes towards the CXCR6 ligand C-X-C motif chemokine ligand 16 (CXCL16) (p<0.05), which is abundantly expressed in the lung. Monocyte CXCR6 and lung CXCL16 were heightened in patients with progressive fibrosing interstitial lung disease (p<0.001), confirming a role for the CXCR6-CXCL16 axis in ongoing lung injury. Conversely, monocytes from long COVID patients with ongoing fatigue exhibited a sustained reduction of the prostaglandin-generating enzyme cyclooxygenase 2 (p<0.01) and CXCR2 expression (p<0.05). These monocyte changes were not present in respiratory syncytial virus or influenza A convalescence. CONCLUSIONS: Our data define unique monocyte signatures that define subgroups of long COVID patients, indicating a key role for monocyte migration in COVID-19 pathophysiology. Targeting these pathways may provide novel therapeutic opportunities in COVID-19 patients with persistent morbidity.

T-Cell Infiltration and Clonality May Identify Distinct Survival Groups in Colorectal Cancer: Development and Validation of a Prognostic Model Based on The Cancer Genome Atlas (TCGA) and Clinical Proteomic Tumor Analysis Consortium (CPTAC).

Predicting the survival outcomes of patients with colorectal cancer (CRC) remains challenging. We investigated the prognostic significance of the transcriptome and tumour-infiltrating lymphocyte T-cell receptor (TIL/Tc-TCR) repertoire and analysed TIL/Tc-TCR sequences of The Cancer Genome Atlas (TCGA) and the Clinical Proteomic Tumor Analysis Consortium (CPTAC) CRC cohorts. Using a multivariate Cox regression, we tested whether TIL/Tc-TCR repertoire, patient and tumour characteristics (stage, sidedness, total non-synonymous mutations, microsatellite instability (MSI) and transcriptional signatures) correlated with patient overall survival (OS) and designed a prognostic nomogram. A multivariate analysis (C-index = 0.75) showed that only patient age, disease stage, TIL/Tc degree of infiltration and clonality were independent prognostic factors for OS. The cut-offs for patients' allocation to TIL/Tc abundance subgroups were determined using a strategy of maximally selected rank statistics with the OptimalCutpoints R package. These were "high", "low" and "very high" (90 th percentile) TIL/Tc infiltration-stratified OS (median not reached, 67 and 44.3 months; p < 0.001); the results were validated in the CPTAC cohort. TIL/Tc clonality was prognostic (median OS in "high" vs. "low" clonality not reached and 67.3 months; p = 0.041) and independent of TIL/Tc infiltration. Whilst tumour sidedness was not prognostic, the "very highly" infiltrated tumours were prevalent among right-sided CRCs (p = 0.039) and showed distinct immunological features, with lower Th1 signature (p = 0.004), higher PD-L1 expression (p < 0.001) and likely enrichment in highly suppressory IL1R1+ Tregs (FoxP3 and IL1R1 overexpression, p < 0.001). TIL/Tc abundance and clonality are independent prognosticators in CRC and, combined with clinical variables, refine risk stratification. We identified a subset of CRCs with "very high" TIL/Tc infiltration, poor prognosis and distinct genetic and immunologic features, which may benefit from alternative therapeutic approaches. These results need validation in prospective patient cohorts.

Interleukin-22 regulates neutrophil recruitment in ulcerative colitis and is associated with resistance to ustekinumab therapy.

The function of interleukin-22 (IL-22) in intestinal barrier homeostasis remains controversial. Here, we map the transcriptional landscape regulated by IL-22 in human colonic epithelial organoids and evaluate the biological, functional and clinical significance of the IL-22 mediated pathways in ulcerative colitis (UC). We show that IL-22 regulated pro-inflammatory pathways are involved in microbial recognition, cancer and immune cell chemotaxis; most prominently those involving CXCR2+ neutrophils. IL-22-mediated transcriptional regulation of CXC-family neutrophil-active chemokine expression is highly conserved across species, is dependent on STAT3 signaling, and is functionally and pathologically important in the recruitment of CXCR2+ neutrophils into colonic tissue. In UC patients, the magnitude of enrichment of the IL-22 regulated transcripts in colonic biopsies correlates with colonic neutrophil infiltration and is enriched in non-responders to ustekinumab therapy. Our data provide further insights into the biology of IL-22 in human disease and highlight its function in the regulation of pathogenic immune pathways, including neutrophil chemotaxis. The transcriptional networks regulated by IL-22 are functionally and clinically important in UC, impacting patient trajectories and responsiveness to biological intervention.

Organoids capture tissue-specific innate lymphoid cell development in mice and humans.

Organoid-based models of murine and human innate lymphoid cell precursor (ILCP) maturation are presented. First, murine intestinal and pulmonary organoids are harnessed to demonstrate that the epithelial niche is sufficient to drive tissue-specific maturation of all innate lymphoid cell (ILC) groups in parallel, without requiring subset-specific cytokine supplementation. Then, more complex human induced pluripotent stem cell (hiPSC)-based gut and lung organoid models are used to demonstrate that human epithelial cells recapitulate maturation of ILC from a stringent systemic human ILCP population, but only when the organoid-associated stromal cells are depleted. These systems offer versatile and reductionist models to dissect the impact of environmental and mucosal niche cues on ILC maturation. In the future, these could provide insight into how ILC activity and development might become dysregulated in chronic inflammatory diseases.

Cyclin-dependent Kinase 9 as a Potential Target for Anti-TNF-resistant Inflammatory Bowel Disease.

BACKGROUND & AIMS: Resistance to single cytokine blockade, namely anti-tumor necrosis factor (TNF) therapy, is a growing concern for patients with inflammatory bowel disease (IBD). The transcription factor T-bet is a critical regulator of intestinal homeostasis, is genetically linked to mucosal inflammation and controls the expression of multiples genes such as the pro-inflammatory cytokines interferon (IFN)-γ and TNF. Inhibiting T-bet may therefore offer a more attractive prospect for treating IBD but remains challenging to target therapeutically. In this study, we evaluate the effect of targeting the transactivation function of T-bet using inhibitors of P-TEFb (CDK9-cyclin T), a transcriptional elongation factor downstream of T-bet. METHODS: Using an adaptive immune-mediated colitis model, human colonic lymphocytes from patients with IBD and multiple large clinical datasets, we investigate the effect of cyclin-dependent kinase 9 (CDK9) inhibitors on cytokine production and gene expression in colonic CD4+ T cells and link these genetic modules to clinical response in patients with IBD. RESULTS: Systemic CDK9 inhibition led to histological improvement of immune-mediated colitis and was associated with targeted suppression of colonic CD4+ T cell-derived IFN-γ and IL-17A. In colonic lymphocytes from patients with IBD, CDK9 inhibition potently repressed genes responsible for pro-inflammatory signalling, and in particular genes regulated by T-bet. Remarkably, CDK9 inhibition targeted genes that were highly expressed in anti-TNF resistant IBD and that predicted non-response to anti-TNF therapy. CONCLUSION: Collectively, our findings reveal CDK9 as a potential target for anti-TNF-resistant IBD, which has the potential for rapid translation to the clinic.

Differential Regulation of Allergic Airway Inflammation by Acetylcholine.

Acetylcholine (ACh) from neuronal and non-neuronal sources plays an important role in the regulation of immune responses and is associated with the development of several disease pathologies. We have previously demonstrated that group 2 innate lymphoid cell (ILC2)-derived ACh is required for optimal type 2 responses to parasitic infection and therefore sought to determine whether this also plays a role in allergic inflammation. RoraCre+ChatLoxP mice (in which ILC2s cannot synthesize ACh) were exposed to an allergenic extract of the fungus Alternaria alternata, and immune responses in the airways and lung tissues were analyzed. Airway neutrophilia and expression of the neutrophil chemoattractants CXCL1 and CXCL2 were enhanced 24 h after exposure, suggesting that ILC2-derived ACh plays a role in limiting excessive pulmonary neutrophilic inflammation. The effect of non-selective depletion of ACh was examined by intranasal administration of a stable parasite-secreted acetylcholinesterase. Depletion of airway ACh in this manner resulted in a more profound enhancement of neutrophilia and chemokine expression, suggesting multiple cellular sources for the release of ACh. In contrast, depletion of ACh inhibited Alternaria-induced activation of ILC2s, suppressing the expression of IL-5, IL-13, and subsequent eosinophilia. Depletion of ACh reduced macrophages with an alternatively activated M2 phenotype and an increase in M1 macrophage marker expression. These data suggest that ACh regulates allergic airway inflammation in several ways, enhancing ILC2-driven eosinophilia but suppressing neutrophilia through reduced chemokine expression.

Heartbreakers or Healers? Innate Lymphoid Cells in Cardiovascular Disease and Obesity.

Cardiovascular diseases (CVDs) are responsible for most pre-mature deaths worldwide, contributing significantly to the global burden of disease and its associated costs to individuals and healthcare systems. Obesity and associated metabolic inflammation underlie development of several major health conditions which act as direct risk factors for development of CVDs. Immune system responses contribute greatly to CVD development and progression, as well as disease resolution. Innate lymphoid cells (ILCs) are a family of helper-like and cytotoxic lymphocytes, typically enriched at barrier sites such as the skin, lung, and gastrointestinal tract. However, recent studies indicate that most solid organs and tissues are home to resident populations of ILCs - including those of the cardiovascular system. Despite their relative rarity, ILCs contribute to many important biological effects during health, whilst promoting inflammatory responses during tissue damage and disease. This mini review will discuss the evidence for pathological and protective roles of ILCs in CVD, and its associated risk factor, obesity.

Transcription factor-driven regulation of ILC1 and ILC3.

Mammalian innate lymphoid cells (ILCs) have functional relevance under both homeostatic and disease settings, such as inflammatory bowel disease (IBD), particularly in the context of maintaining the integrity of mucosal surfaces. Early reports highlighted group 1 and 3 ILC regulatory transcription factors (TFs), T-box expressed in T cells (T-bet; Tbx21) and RAR-related orphan nuclear receptor γt (RORγt; Rorc), as key regulators of ILC biology. Since then, other canonical TFs have been shown to have a role in the development and function of ILC subsets. In this review, we focus on recent insights into the balance between mature ILC1 and ILC3 based on these TFs and how they interact with other key cell-intrinsic molecular pathways. We outline how this TF interplay might be explored to identify novel candidate therapeutic avenues for human diseases.

The TH1 cell lineage-determining transcription factor T-bet suppresses TH2 gene expression by redistributing GATA3 away from TH2 genes.

Lineage-determining transcription factors (LD-TFs) drive the differentiation of progenitor cells into a specific lineage. In CD4+ T cells, T-bet dictates differentiation of the TH1 lineage, whereas GATA3 drives differentiation of the alternative TH2 lineage. However, LD-TFs, including T-bet and GATA3, are frequently co-expressed but how this affects LD-TF function is not known. By expressing T-bet and GATA3 separately or together in mouse T cells, we show that T-bet sequesters GATA3 at its target sites, thereby removing GATA3 from TH2 genes. This redistribution of GATA3 is independent of GATA3 DNA binding activity and is instead mediated by the T-bet DNA binding domain, which interacts with the GATA3 DNA binding domain and changes GATA3's sequence binding preference. This mechanism allows T-bet to drive the TH1 gene expression program in the presence of GATA3. We propose that redistribution of one LD-TF by another may be a common mechanism that could explain how specific cell fate choices can be made even in the presence of other transcription factors driving alternative differentiation pathways.

Mortality risk prediction of high-sensitivity C-reactive protein in suspected acute coronary syndrome: A cohort study.

BACKGROUND: There is limited evidence on the use of high-sensitivity C-reactive protein (hsCRP) as a biomarker for selecting patients for advanced cardiovascular (CV) therapies in the modern era. The prognostic value of mildly elevated hsCRP beyond troponin in a large real-world cohort of unselected patients presenting with suspected acute coronary syndrome (ACS) is unknown. We evaluated whether a mildly elevated hsCRP (up to 15 mg/L) was associated with mortality risk, beyond troponin level, in patients with suspected ACS. METHODS AND FINDINGS: We conducted a retrospective cohort study based on the National Institute for Health Research Health Informatics Collaborative data of 257,948 patients with suspected ACS who had a troponin measured at 5 cardiac centres in the United Kingdom between 2010 and 2017. Patients were divided into 4 hsCRP groups (<2, 2 to 4.9, 5 to 9.9, and 10 to 15 mg/L). The main outcome measure was mortality within 3 years of index presentation. The association between hsCRP levels and all-cause mortality was assessed using multivariable Cox regression analysis adjusted for age, sex, haemoglobin, white cell count (WCC), platelet count, creatinine, and troponin. Following the exclusion criteria, there were 102,337 patients included in the analysis (hsCRP <2 mg/L (n = 38,390), 2 to 4.9 mg/L (n = 27,397), 5 to 9.9 mg/L (n = 26,957), and 10 to 15 mg/L (n = 9,593)). On multivariable Cox regression analysis, there was a positive and graded relationship between hsCRP level and mortality at baseline, which remained at 3 years (hazard ratio (HR) (95% CI) of 1.32 (1.18 to 1.48) for those with hsCRP 2.0 to 4.9 mg/L and 1.40 (1.26 to 1.57) and 2.00 (1.75 to 2.28) for those with hsCRP 5 to 9.9 mg/L and 10 to 15 mg/L, respectively. This relationship was independent of troponin in all suspected ACS patients and was further verified in those who were confirmed to have an ACS diagnosis by clinical coding. The main limitation of our study is that we did not have data on underlying cause of death; however, the exclusion of those with abnormal WCC or hsCRP levels >15 mg/L makes it unlikely that sepsis was a major contributor. CONCLUSIONS: These multicentre, real-world data from a large cohort of patients with suspected ACS suggest that mildly elevated hsCRP (up to 15 mg/L) may be a clinically meaningful prognostic marker beyond troponin and point to its potential utility in selecting patients for novel treatments targeting inflammation. TRIAL REGISTRATION: ClinicalTrials.gov - NCT03507309.

A population of naive-like CD4+ T cells stably polarized to the TH 1 lineage.

T-bet is the lineage-specifying transcription factor for CD4+ TH 1 cells. T-bet has also been found in other CD4+ T cell subsets, including TH 17 cells and Treg, where it modulates their functional characteristics. However, we lack information on when and where T-bet is expressed during T cell differentiation and how this impacts T cell differentiation and function. To address this, we traced the ontogeny of T-bet-expressing cells using a fluorescent fate-mapping mouse line. We demonstrate that T-bet is expressed in a subset of CD4+ T cells that have naïve cell surface markers and transcriptional profile and that this novel cell population is phenotypically and functionally distinct from previously described populations of naïve and memory CD4+ T cells. Naïve-like T-bet-experienced cells are polarized to the TH 1 lineage, predisposed to produce IFN-γ upon cell activation, and resist repolarization to other lineages in vitro and in vivo. These results demonstrate that lineage-specifying factors can polarize T cells in the absence of canonical markers of T cell activation and that this has an impact on the subsequent T-helper response.

Sustained Post-Developmental T-Bet Expression Is Critical for the Maintenance of Type One Innate Lymphoid Cells In Vivo.

Innate lymphoid cells (ILC) play a significant role in the intestinal immune response and T-bet+ CD127+ group 1 cells (ILC1) have been linked to the pathogenesis of human inflammatory bowel disease (IBD). However, the functional importance of ILC1 in the context of an intact adaptive immune response has been controversial. In this report we demonstrate that induced depletion of T-bet using a Rosa26-Cre-ERT2 model resulted in the loss of intestinal ILC1, pointing to a post-developmental requirement of T-bet expression for these cells. In contrast, neither colonic lamina propria (cLP) ILC2 nor cLP ILC3 abundance were altered upon induced deletion of T-bet. Mechanistically, we report that STAT1 or STAT4 are not required for intestinal ILC1 development and maintenance. Mice with induced deletion of T-bet and subsequent loss of ILC1 were protected from the induction of severe colitis in vivo. Hence, this study provides support for the clinical development of an IBD treatment based on ILC1 depletion via targeting T-bet or its downstream transcriptional targets.

The Th1 cell regulatory circuitry is largely conserved between human and mouse.

Gene expression programs controlled by lineage-determining transcription factors are often conserved between species. However, infectious diseases have exerted profound evolutionary pressure, and therefore the genes regulated by immune-specific transcription factors might be expected to exhibit greater divergence. T-bet (Tbx21) is the immune-specific, lineage-specifying transcription factor for T helper type I (Th1) immunity, which is fundamental for the immune response to intracellular pathogens but also underlies inflammatory diseases. We compared T-bet genomic targets between mouse and human CD4+ T cells and correlated T-bet binding patterns with species-specific gene expression. Remarkably, we found that the majority of T-bet target genes are conserved between mouse and human, either via preservation of binding sites or via alternative binding sites associated with transposon-linked insertion. Species-specific T-bet binding was associated with differences in transcription factor-binding motifs and species-specific expression of associated genes. These results provide a genome-wide cross-species comparison of Th1 gene regulation that will enable more accurate translation of genetic targets and therapeutics from pre-clinical models of inflammatory and infectious diseases and cancer into human clinical trials.

A Crohn's Disease-associated IL2RA Enhancer Variant Determines the Balance of T Cell Immunity by Regulating Responsiveness to IL-2 Signalling.

BACKGROUND AND AIMS: Differential responsiveness to interleukin [IL]-2 between effector CD4+ T cells [Teff] and regulatory T cells [Treg] is a fundamental mechanism of immunoregulation. The single nucleotide polymorphism [SNP] rs61839660, located within IL2RA [CD25], has been associated with the development of Crohn's disease [CD]. We sought to identify the T cell immune phenotype of IBD patients who carry this SNP. METHODS: Teff and Treg were isolated from individuals homozygous [TT], heterozygous [CT], or wild-type [CC] for the minor allele at rs61839660, and used for phenotyping [flow cytometry, Cytometry Time Of Flight] functional assays or T cell receptor [TCR] sequencing. Phosphorylation of signal transducer and activator of transcription 5 [STAT5] was assessed in response to IL-2, IL-7, and in the presence of basiliximab, a monoclonal antibody directed against CD25. Teff pro-inflammatory cytokine expression levels were assessed by reverse transcription quantitative polymerase chain reaction after IL-2 and/or TCR stimulation. RESULTS: Presence of the minor T allele enhances CD25 expression, leading to increased STAT5 phosphorylation and pro-inflammatory cytokine transcript expression by Teff in response to IL-2 stimulation in vitro. Teff from TT individuals demonstrate a more activated gut homing phenotype. TCR sequencing analysis suggests that TT patients may have a reduced clonal capacity to mount an optimal regulatory T cell response. CONCLUSIONS: rs61839660 regulates the responsiveness of T cells to IL-2 signalling by modulating CD25 expression. As low-dose IL-2 is being trialled as a selective Treg modulator in CD, these findings highlight the potential for adverse effects in patients with this genotype.

MicroRNA-142 Critically Regulates Group 2 Innate Lymphoid Cell Homeostasis and Function.

Innate lymphoid cells are central to the regulation of immunity at mucosal barrier sites, with group 2 innate lymphoid cells (ILC2s) being particularly important in type 2 immunity. In this study, we demonstrate that microRNA(miR)-142 plays a critical, cell-intrinsic role in the homeostasis and function of ILC2s. Mice deficient for miR-142 expression demonstrate an ILC2 progenitor-biased development in the bone marrow, and along with peripheral ILC2s at mucosal sites, these cells display a greatly altered phenotype based on surface marker expression. ILC2 proliferative and effector functions are severely dysfunctional following Nippostrongylus brasiliensis infection, revealing a critical role for miR-142 isoforms in ILC2-mediated immune responses. Mechanistically, Socs1 and Gfi1 expression are regulated by miR-142 isoforms in ILC2s, impacting ILC2 phenotypes as well as the proliferative and effector capacity of these cells. The identification of these novel pathways opens potential new avenues to modulate ILC2-dependent immune functions.

An update on the roles of immune system-derived microRNAs in cardiovascular diseases.

Cardiovascular diseases (CVD) are a leading cause of human death worldwide. Over the past two decades, the emerging field of cardioimmunology has demonstrated how cells of the immune system play vital roles in the pathogenesis of CVD. MicroRNAs (miRNAs) are critical regulators of cellular identity and function. Cell-intrinsic, as well as cell-extrinsic, roles of immune and inflammatory cell-derived miRNAs have been, and continue to be, extensively studied. Several 'immuno-miRNAs' appear to be specifically expressed or demonstrate greatly enriched expression within leucocytes. Identification of miRNAs as critical regulators of immune system signalling pathways has posed the question of whether and how targeting these molecules therapeutically, may afford opportunities for disease treatment and/or management. As the field of cardioimmunology rapidly continues to advance, this review discusses findings from recent human and murine studies which contribute to our understanding of how leucocytes of innate and adaptive immunity are regulated-and may also regulate other cell types, via the actions of the miRNAs they express, in the context of CVD. Finally, we focus on available information regarding miRNA regulation of regulatory T cells and argue that targeted manipulation of miRNA regulated pathways in these cells may hold therapeutic promise for the treatment of CVD and associated risk factors.

ILC1 drive intestinal epithelial and matrix remodelling.

Organoids can shed light on the dynamic interplay between complex tissues and rare cell types within a controlled microenvironment. Here, we develop gut organoid cocultures with type-1 innate lymphoid cells (ILC1) to dissect the impact of their accumulation in inflamed intestines. We demonstrate that murine and human ILC1 secrete transforming growth factor ß1, driving expansion of CD44v6+ epithelial crypts. ILC1 additionally express MMP9 and drive gene signatures indicative of extracellular matrix remodelling. We therefore encapsulated human epithelial-mesenchymal intestinal organoids in MMP-sensitive, synthetic hydrogels designed to form efficient networks at low polymer concentrations. Harnessing this defined system, we demonstrate that ILC1 drive matrix softening and stiffening, which we suggest occurs through balanced matrix degradation and deposition. Our platform enabled us to elucidate previously undescribed interactions between ILC1 and their microenvironment, which suggest that they may exacerbate fibrosis and tumour growth when enriched in inflamed patient tissues.