Fibrosis, a common end point of chronic inflammatory diseases across organs, remains a major therapeutic challenge. Macrophages are central regulators of fibrotic remodeling, with evidence suggesting their direct role through macrophage-to-myofibroblast transition. However, the upstream mechanisms governing macrophage fibrogenic reprogramming remain unclear. Using a laser-induced mouse model of subretinal fibrosis, whether adhesion-dependent mechanotransduction regulates macrophage-driven fibrosis was investigated. Bulk RNA sequencing of retinal pigment epithelium-choroid tissues revealed significant enrichment of adhesion-related genes and focal adhesion pathways, with up-regulation of integrins such as Itgb2, Itgal, and Itgax. Increased integrin expression and focal adhesion kinase (FAK) phosphorylation were detected in infiltrating F4/80+ macrophages within fibrotic lesions. Bone marrow-derived macrophages under high adherence conditions expressed higher levels of fibrosis-related genes and FAK-related genes such as Ptk2 and Ptk2b. FAK inhibitor PF562271 suppressed transforming growth factor-β1-induced up-regulation of fibrosis-related genes (Col1a1, Fn1, and Acta2) and α-smooth muscle actin in macrophages. I.P. administration of PF562271 reduced expression of adhesion molecules ITGB2 and ITGAL on circulating monocytes and alleviated subretinal fibrosis. These findings identify integrin-FAK-mediated mechanotransduction as a key regulator of macrophage fibrogenic reprogramming and macrophage-to-myofibroblast transition. Adhesion-dependent signaling is highlighted as a conserved pathway linking tissue remodeling to fibrotic macrophage activation and FAK as a potential therapeutic target.

The gastrointestinal tract is densely innervated by the enteric nervous system (ENS), a complex neural network that regulates intestinal physiology. Emerging advances highlight the essential contributions of ENS to immune homeostasis and inflammatory responses within the gut. This review synthesizes current understanding of the interactions between the intrinsic enteric neurons and various intestinal immune cells, epithelium cells and the microbiome. We also discuss recent technological developments that enhance our ability to dissect the immunomodulatory functions of enteric neurons. Elucidating these complex communication pathways is critical for advancing our understanding of gut function and mucosal inflammation, and for developing novel therapeutic strategies for gastrointestinal disorders.

Germinal center B cell responses are defined by many positive regulators of affinity maturation, but few components that restrain clonal dominance, notably Nr4a1, are known. We reveal an unsuspected role for BLIMP1 (Prdm1)-a plasma cell determinant-as a feedback regulator of affinity maturation. Single-cell RNA and B cell receptor (BCR) sequencing showed that B cell-specific Prdm1 loss drives an exaggerated germinal center reaction with larger clones, increased somatic hypermutation and greater clonal dominance, independent of Nr4a1. Single-cell chromatin profiling with base-resolution modeling indicated that Blimp-1 represses expression of BCR-signaling genes, gating chromatin accessibility at interferon-stimulated response elements, Ets-interferon regulatory factor composite elements, nuclear factor kappa B and Oct motifs. In the absence of BLIMP1, enhanced BCR-signaling augments activities of transcription factors that promote G1-S transition during light zone (LZ) selection and fuel dark zone (DZ) expansion. Thus, BLIMP1 attenuates BCR signaling and constrains the LZ to DZ transition, fine-tuning clonal competition, thereby maintaining repertoire diversity.

Pathologic myopia is a major cause of irreversible visual impairment worldwide and is characterized by excessive axial elongation accompanied by progressive retinal degeneration. Whether vision loss results primarily from passive retinal stretching or selective neurodegeneration remains unclear, hindering the development of effective neuroprotective and regenerative therapies. Here, we investigated retinal neuronal, vascular, and glial alterations in retinal pigment epithelium (RPE)-specific Lrp2 knockout (Best1-Cre/Lrp2fl/fl conditional knockout, CKO) model of pathologic myopia. The CKO mice were examined longitudinally using multimodal ocular imaging, electroretinography, optokinetic testing, fluorescein angiography, and quantitative immunohistochemistry analysis. CKO phenotype+ mice developed early-onset, progressive axial elongation and high myopia, accompanied by fundus features closely resembling human pathologic myopia, including peripapillary and patchy chorioretinal atrophy. Retinal function was markedly impaired, with significant reductions in scotopic a-, b-, and c-wave amplitudes. Although axial elongation resulted in a 1.98-fold increase in retinal surface area and a 55.95% reduction in retinal thickness, quantitative correction for retinal expansion revealed selective neuronal loss rather than uniform retinal degeneration. Total numbers of rods, cones, horizontal cells, and GABAergic amacrine cells were reduced by 22, 40, 30, and 57%, respectively, together with a 66% loss of photoreceptor synaptic ribbons. In contrast, retinal ganglion cells and bipolar cells exhibited reduced density but preserved absolute cell numbers. These neuronal changes were accompanied by retinal and choroidal microvascular degeneration, Müller gliosis, microglial activation and subretinal accumulation, and RPE dysmorphology. Our findings demonstrate that axial elongation induces neuron subtype–specific degeneration rather than generalized retinal thinning. Our study identifies photoreceptors, horizontal cells, and inhibitory amacrine cells as particularly vulnerable populations and implicates impaired RPE support, neurovascular dysfunction, and chronic glial activation as key mechanisms driving myopic retinopathy. This study provides a mechanistic framework for developing targeted neuroprotective and regeneration-based therapies for pathologic myopia.

Abstract Background Subretinal fibrosis causes irreversible vision loss in neovascular age-related macular degeneration (nAMD). Sustained macular inflammation drives the initiation and progression of fibrosis by activating profibrotic cells and perpetuating tissue damage. This study investigated the therapeutic potential of mesenchymal stem cell-derived extracellular vesicles (MSC-EVs) in mitigating nAMD-associated subretinal fibrosis. Methods MSC-EVs were prepared from human bone marrow-derived MSCs and characterized using nanoparticle tracking analysis, transmission electron microscopy, and Western Blotting. Subretinal fibrosis was induced in C57BL/6J mice using the two-stage laser-induced model. MSC-EVs were injected either intravitreally (1 × 10 8 particles/eye, single injection) or retro-orbitally (1 × 10 8 particles, two injections four days apart) immediately after the second laser. Eyes were collected 10 days post-second laser for immunostaining of collagen-1 and CD31 or iso-lectin B4. In vitro, primary human RPE and ARPE-19 cells were treated with TGF-β2 (10 ng/mL) to induce epithelial-mesenchymal transition (EMT); peritoneal macrophages were treated with TGF-β1 (10 ng/mL) to induce macrophage-to-myofibroblast transition (MMT). After 48 h, cells were treated with MSC-EVs (cell-to-MSC-EV ratio = 1:2000) for 3 days. Myofibroblast markers (αSMA, fibronectin, and collagen-1) were examined by immunocytochemistry and quantitative PCR (qPCR). Human iPCS-derived macrophages (iMACs), bone-marrow-derived macrophages, peritoneal macrophages, and BV2 microglia were treated with LPS (100 ng/mL) and IFN-γ (20 ng/mL) for 24 h with or without MSC-EVs (1:2000). Small RNA sequencing was used to identify specific functional molecules within MSC-EVs. Immune-related gene expressions were evaluated by qPCR. Results Intravitreal and retroorbital administration of MSC-EVs reduced collagen-1 + fibrotic lesions by 46% and 30%, respectively, and significantly inhibited infiltrating Iba-1 + cells. In vitro, MSC-EVs attenuated TGF-β2-induced upregulation of αSMA, fibronectin, and collagen-1 at both protein and mRNA levels in RPE cells. Similarly, the expression of Acta2 , Fn1 , and Col1a1 in TGF-β1-treated macrophages was also significantly reduced following MSC-EV treatment. In LPS + IFN-γ-stimulated immune cells, MSC-EVs significantly suppressed the expression of Il6 and Il1b in all cell types, and reduced the expression of Inos , Tnfa , and Cd86 in iMACs, peritoneal macrophages, and BV2 cells. Enriched hsa-miR-21-5p was identified in MSC-EVs and involved in the TGF-β-related signaling pathway. Overexpression of miR-21-5p mimic abrogated the TGF-β1-driven upregulation of pro-fibrotic markers in RPE and macrophages. Conclusions Local administration of MSC-EVs effectively mitigated subretinal fibrosis and reduced inflammation in the mouse model of nAMD, potentially via miR-21-5p-mediated attenuation of EMT and MMT, and suppression of inflammation. MSC-EVs represent a novel cell-free therapeutic strategy for macular fibrosis in nAMD.

Macular subretinal fibrosis represents the end-stage complication of neovascular age-related macular degeneration (nAMD). Despite the widespread use of anti-VEGF therapy, between 20 and 67% of nAMD patients develop this sight-threatening condition within 2 to 7 years, making fibrosis one of the main reasons responsible for anti-VEGF resistance and irreversible visual loss. Unlike the quiescent scar of a healed skin wound, subretinal fibrosis is a dynamic, vascularised lesion characterised by persistent and active inflammation, a process aptly termed "hot fibrosis". The molecular mechanisms that drive the conversion of neovascularisation into fibrovascular membrane remain poorly defined. This review summarises the clinical features of macular fibrosis in nAMD. It will then detail the pathological insights gained from advanced clinical high-resolution imaging and post-mortem histopathological investigations. We will also explore how advances in our understanding of the pathogenesis of macular fibrosis, as revealed by preclinical models, are uncovering novel therapeutic targets. Promising approaches include dual-targeting agents (e.g., inhibiting VEGF and Ang-2 or complement pathways), as well as repurposing FDA-approved anti-fibrotics, such as pirfenidone or nintedanib, for ocular delivery. The repurposing of drugs with established safety profiles, reformulated for local intraocular use, offers a particularly resource-efficient pathway.

To investigate the stage-specific role of NK1.1⁺ cells in two experimental models of retinal injury: laser-induced choroidal neovascularization (CNV) and retinal detachment (RD). The study used C57BL/6J mice (8–12 weeks). NK1.1+ cells were depleted either during the acute phase (< 48 h after injury) or throughout the disease progression (i.e., 2, 4, and 6 days after laser injury) using anti-NK1.1 antibody (PK136). The outcome measurements included CNV lesion size in RPE/choroid/sclera flatmounts (following collagen IV staining), photoreceptor death (with TUNEL staining), RPE/choroidal infiltrating myeloid cells (by Iba-1 immunostaining), and innate lymphocytes (e.g., assessed in blood, spleen, and ocular single-cell suspension by flow cytometry and immunofluorescence on RPE/choroid/sclera for CD45NK1.1⁺ (including NK and ILC1), CD45+CD3⁻T-bet⁺ (ILC1), CD45+CD127+GATA3+ (ILC2), and CD45+CD127+RORγ+ (ILC3)). Retinal laser injury increased circulating NK, ILC1, and ILC2 cells at 1 h (p < 0.05), accompanied by the accumulation of infiltrating ILC1/2 cells (p < 0.001). The number of infiltrating NK1.1+ cells increased progressively from 1 to 24 h (CNV, p < 0.001; RD, p < 0.01). Sustained NK1.1+ cell depletion significantly reduced circulating NK and ILC1 cells, exacerbated collagen IV+ CNV (p < 0.05) and increased the number of infiltrating Iba-1+ cells (p < 0.001). In contrast, NK1.1⁺ cell (NK and ILC1) depletion either immediately after or 48 h before injury significantly reduced the severity of laser-induced CNV (p < 0.05) and suppressed Iba-1+ cell infiltration (p < 0.05). Early NK1.1+ cell (NK and ILC1) depletion also attenuated RD-mediated TUNEL+ apoptotic photoreceptors (p < 0.01) and reduced infiltrating Iba-1+ cells (p < 0.05). NK1.1⁺ cells exert stage-dependent effects in retinal injury, amplifying inflammation during the acute phase while contributing to immune regulation at later stages.

Purpose:Subretinal fibrosis is a major cause of irreversible vision loss in neovascular age-related macular degeneration (AMD), yet no effective antifibrotic therapies exist due to poorly defined molecular drivers. This study aimed to derive and validate a persistent ocular fibrosis signature using the two-stage laser-induced subretinal fibrosis mouse model and to assess its translational relevance in human AMD. Methods:RNA sequencing (RNA-seq) was performed on retinal pigment epithelium (RPE)/choroid tissues collected at days 3, 6, and 10 following the second laser injury in the two-stage model. A novel core fibrosis signature of 88 persistently upregulated genes was observed across all time points and cross-validated in an independent mouse dataset (GSE189555) and multiple human AMD RNA-seq datasets (GSE115828, GSE146887, GSE135092), including surgically extracted choroidal neovascularization (CNV) membranes and macular RPE/choroid samples. Key signature genes were further validated by immunofluorescence in human subretinal fibrotic and age-matched tissues. Results:The novel ocular fibrosis signature was enriched in epithelial-mesenchymal transition, complement activation, and inflammatory pathways and showed choroid-specific expression with minimal retinal involvement. Cross-validation on independent chronic mouse data and multiple human AMD datasets (peripheral retina, surgically extracted CNV membranes, and macular RPE/choroid) confirmed a progressive enrichment in advanced disease stages. Immunofluorescence in human fibrotic tissue validated key genes (tenascin C, tissue inhibitor of metalloproteinases 1, and apelin receptor) and showed colocalization with myofibroblast-like cells and microglia. Conclusions:We identified a novel, persistent, choroid-specific ocular fibrosis signature with strong cross-species conservation, highlighting fibrogenic-associated drivers and providing a valuable translational tool to understand subretinal fibrosis development and targeted antifibrotic therapies in neovascular AMD.

Crohn's disease (CD) is a prevalent type of inflammatory bowel disease (IBD) with dysregulated antibody responses. However, there is a lack of comprehensive analysis of B cell responses in CD. Here, we collected B cells from the small intestine, colon and blood of CD patients and control subjects. Through the coupled analysis of transcriptome and immunoglobulin (Ig) gene in individual cells, we characterized the cellular composition, transcriptome and Ig clonotype in different B cell subtypes. We observed shared disruptions in plasma cell (PC) responses between different IBD subtypes. We revealed heterogeneity in memory B cells (MBCs) and showed a positive correlation between gut resident-like MBCs and disease severity. Furthermore, our clonotype analysis demonstrated an increased direct differentiation of MBCs into PCs in CD patients. Overall, this study demonstrates significantly altered B cell responses associated with chronic inflammation during CD and highlights the potential role of mucosal MBCs in CD pathogenesis.

Pathological fibrosis is a chronic process characterized by excessive deposition of extracellular matrix (ECM), which disrupts tissue architecture and function and accelerates disease progression in organs such as the lungs, liver, heart, kidneys, skin, and eyes. Recent studies have identified leucine-rich α-2 glycoprotein-1 (LRG1), a secreted glycoprotein known for modulating angiogenesis and immune responses, as a key player in the pathogenesis of fibrotic disorders. LRG1 expression is up-regulated by multiple pro-fibrotic mediators, including transforming growth factor-beta (TGF-β), and has been shown to regulate fibroblast differentiation, myofibroblast activation, and ECM production. Through interactions with the TGF-β signaling pathway and other cascades, LRG1 plays a crucial role in pathological fibrosis. Elucidating the molecular mechanisms by which LRG1 drives fibrogenic responses will pave the way for novel therapeutic strategies targeting pathological fibrosis. This review explores the emerging functions of LRG1 in fibrosis of different organs and discusses therapeutic approaches aimed at mitigating fibrosis through LRG1 inhibition.

Purpose:The purpose of this study was to understand how the gut microbial system responds to retinal injury. Methods:Adult C57BL/6J mice were subjected to retinal laser burns or hypotony-induced retinal detachment (RD). One, 4, and 24 hours later, gut permeability (8 male mice and 8 female mice) was assessed using Evan's blue assay and the expression of ZO-1 in intestinal epithelial cells was examined by immunofluorescence. Circulating immune cells were evaluated by flow cytometry. The feces from control and lasered mice (n = 8) were collected under strict sterile conditions and processed for 16S DNA paired-end sequencing using the Illumina platform. The impact of gut dysbiosis on retinal wound healing was evaluated following treatment with Peros antibiotics (n = 8). Retinal pathologies were examined by immunohistochemistry. Results:Retinal laser injury significantly altered gut microbial profiles within 1 hour (β-diversity, multi-response permutation procedure [MRPP], P = 0.05). The abundance of Lignipirellula and Faecalibacterium was 100- and 6.67-fold lower, and the abundance of Akkermansia and Colidextribacter was 3.65- and 17.72-fold higher than non-lasered controls, respectively. Retinal laser burns and RD, not sham surgery, increased gut permeability at 1 hour and 4 hours by 3.82- and 24.76-fold, respectively, disrupted intestinal epithelial ZO-1 expression, accompanied by an increased population of circulating neutrophils and monocytes (P < 0.01) at 1 hour and 4 hours. Antibiotic treatment attenuated laser-/RD-induced gut permeability and the increased neutrophils and monocytes (in RD, P < 0.05). Antibiotic treatment also significantly reduced the severity of laser-induced choroidal neovascularization (CNV; P < 0.001) and RD-mediated photoreceptor apoptosis (P < 0.01), and suppressed Gr-1+ neutrophils (CNV, P < 0.001) and Iba-1+ cell infiltration (P < 0.001). Conclusions:A retina-gut axis exists. Retinal injury induces rapid gut microbial alteration, which in turn modulates innate immune cell activation and regulates the wound healing response.

Age-related macular degeneration (AMD) is a prevalent neuroinflammation condition and the leading cause of irreversible blindness among the elderly population. Smoking significantly increases AMD risk, yet the mechanisms remain unclear. Here, we investigate the role of Sema4D-PlexinB1 axis in the progression of AMD, in which Sema4D-PlexinB1 is highly activated by smoking. Using patient-derived samples and mouse models, we discover that smoking increases the presence of Sema4D on the surface of CD8+ T cells that migrate into the choroidal neovascularization (CNV) lesion via CXCL12-CXCR4 axis and interact with its receptor PlexinB1 on choroidal pericytes. This leads to ROR2-mediated PlexinB1 phosphorylation and pericyte activation, thereby disrupting vascular homeostasis and promoting neovascularization. Inhibition of Sema4D reduces CNV and improves the benefit of anti-VEGF treatment. In conclusion, this study unveils the molecular mechanisms through which smoking exacerbates AMD pathology, and presents a potential therapeutic strategy by targeting Sema4D to augment current AMD treatments.

Intrinsic enteric neurons (iENs) form a crucial neuronal network within the myenteric and submucosal plexus of the gastrointestinal tract, primarily responsible for regulating gut peristalsis. The mechanisms by which iENs sense and integrate dietary and microbial signals to regulate intestinal homeostasis and inflammation remain unclear. Here, we showed that environmental sensor aryl hydrocarbon receptor (AHR) was expressed in different iEN subsets in the ileum and colon and that AHR ligands differentially modulated iEN activity in these regions. Genetic perturbation of Ahr in neurons increased iEN activation in the ileum but, conversely, decreased it in the colon in response to different intestinal pathogens. Furthermore, neuronal AHR deficiency enhanced the clearance of bacterial pathogens, which was associated with increased proliferation and abundance of group 3 innate lymphoid cells in the ileum. Together, our findings demonstrate the region-specific functions of AHR in neurons in response to infections.

Ocular fibrosis, a severe consequence of excessive retinal wound healing, can lead to vision loss following retinal injury. Proliferative vitreoretinopathy (PVR), a common form of ocular fibrosis, is a major cause of blindness, characterized by the formation of extensive fibrous proliferative membranes. Understanding the cellular origins of PVR-associated fibroblasts (PAFs) is essential to decipher the mechanisms of ocular wound healing. In this study, we combined single-cell transcriptomics with genetic lineage tracing to map the contributions of retinal pigment epithelial (RPE) cells, immune cells, and Müller cells to disease progression. RPE cells were found to constitute the largest fraction of cells within PVR lesions, transitioning through metabolic, proliferative, and epithelial-to-mesenchymal transition stages during their conversion to PAFs. These cells exhibited remarkable plasticity and heterogeneity. Notably, Pdgfrb + RPE cells demonstrated significant morphological plasticity, transitioning toward a fibroblast-like phenotype, while macrophage-like RPE cells acquired inflammation-related functions post-PVR. Cell communication network analysis identified Thbs1 (encoding TSP-1) as a key hub gene driving RPE cell fate transitions during PVR. Importantly, therapeutic antibodies targeting TSP-1 significantly mitigated PVR progression. This study provides a detailed roadmap of fibrosis formation during ocular wound healing and highlights the therapeutic potential of targeting TSP-1 in the management of PVR.

Purpose:To investigate the differential role of infiltrating CCR2+ macrophages and CX3CR1+ microglia in neovascular AMD (nAMD)-mediated subretinal fibrosis. Methods:Subretinal fibrosis was induced using the two-stage laser protocol in C57BL/6J or CX3CR1gfp/+ mice. The fibrotic lesion was detected using collagen-1 staining in retinal pigment epithelial /choroidal flatmounts. Infiltrating macrophages and microglial were identified using F4/80, CCR2, and CX3CR1 markers at one, three, six, and 10 days after the second laser. Circulating CCR2+ monocytes were depleted using the MC-21 antibody, whereas CX3CR1+ microglia were depleted using PLX5622. BV2 microglia were treated with TGF-β1 for 96 hours, and their profibrotic potential was examined by quantitative PCR and immunocytochemistry. Results:Subretinal fibrosis lesions developed three days after the second laser, accompanied by persistent CCR2+F4/80+ macrophage and CX3CR1+ cell infiltration. Inflammation in the first three days after the second laser was dominated by filtrating CX3CR1+ cells, and the number increased until day (D) 10 post-second laser. Depletion of CCR2+ monocytes from D5-10 significantly reduced the vascular and fibrotic components of the lesion, while CX3CR1+ cell depletion reduced Isolectin B4+ but not collagen-1+ lesion size. Bone marrow-derived macrophages from D6 and D10 mice expressed significantly higher levels of α-smooth muscle actin (α-SMA) and collagen-1 compared to cells from D1 and D3. TGFβ1 treatment increased TMEM119, CX3CR1, IL1b and iNOS gene expression but did not affect Acta2 and Col1a1 gene expression in BV2 cells. Conclusions:CCR2+ monocytes, but not CX3CR1+ microglia, critically contribute to the development of subretinal fibrosis in nAMD.

Purpose:Retinal vein occlusion (RVO) is a common retinal vascular disease that severely threatens visual function. This study aims to elucidate the role of the complement C3/C3aR signaling pathway in a laser-induced RVO mouse model and to explore its potential as a therapeutic target. Methods:RVO was induced in C57BL/6J mice using laser photocoagulation combined with photosensitizer dye administration. Two days later, retinal tissues were collected for bulk RNA sequencing. The activation of the C3/C3aR signaling pathway was validated through RT-qPCR and Western blot. The C3aR antagonist SB290157 (C3aRA) was administered intravitreally and retinal morphological and functional changes were examined 1, 2, and 8 days later by optical coherence tomography (OCT), fundus photography (FP), and fluorescein angiography (FA), optomotor response (OKR) test, and electroretinogram (ERG). Results:RVO mice exhibited marked increases in retinal thickness (P < 0.001) and fluorescence leakage (P < 0.01) compared to the sham-laser group. Bulk RNA-seq revealed significant upregulation of the complement pathway. Elevated expression of C3 and C3aR (P < 0.05) was confirmed by RT-qPCR and Western blot. Blocking C3aR with SB290157 significantly alleviated RVO-induced retinal edema, vascular leakage, and structural damage. Functional assessment showed that SB290157 treatment significantly improved contrast sensitivity (P < 0.05), increased b-wave (P < 0.001), and oscillatory potentials (Ops) amplitudes (P < 0.05) in RVO mice. RNA-seq analysis demonstrated that SB290157 significantly reduced the inflammatory mediator-related pathways and upregulated visual perception pathways (P < 0.05). Conclusions:The complement C3/C3aR signaling pathway is critically involved in RVO-induced retinal damage and targeting this pathway may be a promising approach for RVO treatment.

Tissue function in homeostasis and disease arise from coordinated interactions between different cell types and can vary between individuals in a population, in part due to the impact of genetic variants. In barrier tissues such as the gut, innate lymphoid cells (ILCs) play critical roles in maintaining tissue homeostasis and immunity, but the genetic basis and associated regulatory circuitry affecting tissue ILCs remain largely unknown. Here, we systematically mapped the regulatory functions of genetic variants across 273,370 gut ILCs profiled by single cell RNA-seq from 274 diversity outbred (DO) mice. Computational analysis identified quantitative trait loci (QTL) impacting ILCs at three levels: cell subtype-specific mRNA expression (local and distal), cell subset proportions, and ILC gene expression programs (in trans ), of which we experimentally validated a pivotal role for the transcription factor Rbpj in regulating ILC3s. All three classes of ILC QTL display polygenic inheritance signatures and broadly overlap with QTL affecting peripheral cytokine levels that we measured in another 261 DO mice. Strikingly, nearly half of trans -QTL affecting ILC traits did not overlap (within a 50-kb window) genes expressed in ILCs, and such loci were enriched for genes expressed by non-ILCs especially enteric neurons and intestinal glia. This suggests that such loci may act instead across cell type boundaries and allowed us to recover causal trans -cell type regulatory circuits in the tissue, including loci encoding multiple neuron-expressed peptides, epithelial-cell expressed Sox9 , and phagocyte-expressed Ccl17 . Finally, human orthologs of genes in QTL impacting gene expression and cell proportions are enriched for autoimmune disease risk heritability, suggesting their relevance to human disease. Our findings highlight how the impact of genetic variants may propagate to maintain tissue homeostasis and show a path to understand causality in tissue biology using quantitative genetics. ### Competing Interest Statement The authors declare competing financial interests: A.R. is a cofounder and equity holder of Celsius Therapeutics, an equity holder in Immunitas, and was an SAB member of ThermoFisher Scientific, Syros Pharmaceuticals, Neogene Therapeutics and Asimov until July 31, 2020. From August 1, 2020, A.R. is an employee of Genentech. R.J.X. is a cofounder of Celsius Therapeutics, Jnana Therapeutics, and a member of the scientific advisory board of Magnet Bio medicine, Nestle and Moonlake Immunotherapeutics.

Emerging studies reveal that neurotransmitters and neuropeptides play critical roles in regulating anti-helminth immune responses, hinting at the potential of intrinsic enteric neurons (iENs) in orchestrating intestinal immunity. Whether and how iENs are activated during infection and the potential neuroimmune interactions involved remain poorly defined. Here, we found that helminth infection activated a subset of iENs. Single-nucleus RNA sequencing (snRNA-seq) of iENs revealed alterations in the transcriptional profile of interleukin (IL)-13R+ intrinsic primary afferent neurons (IPANs), including the upregulation of the neuropeptide β-calcitonin gene-related peptide (CGRP). Using genetic mouse models and engineered viral tools, we demonstrated that group 2 innate lymphoid cell (ILC2)-derived IL-13 was required to activate iENs via the IL-13R, leading to iEN production of β-CGRP, which subsequently inhibited ILC2 responses and anti-helminth immunity. Together, these results reveal a previously unrecognized bi-directional neuroimmune crosstalk in the intestine between a subset of iENs and ILC2s, which influences pathogen clearance.

Age-related retinal degeneration, such as diabetic retinopathy and age-related macular degeneration, are major causes of blindness in modern society. Recent studies suggest that dysbiosis and intraocular translocation of bacteria from the blood circulation are critically involved in retinal degeneration. We hypothesise that the blood-retinal barrier (BRB) cells can protect the neuroretina from blood-borne pathogens by producing antimicrobial peptides (AMPs). The antimicrobial activity may decline during ageing, putting the retina at risk of low-degree chronic inflammation and degeneration. Here, we found that the retinal pigment epithelial (RPE) cells, which form the outer BRB, express a variety of AMPs/AMP precursors, including APP, RARRES2, FAM3A, HAMP, CAMP, GNLY, and PI3. Senescent RPE cells expressed lower levels of APP and RARRES2 mRNA, accompanied by increased intracellular retention of E. coli in a bactericidal assay. Silencing APP, not RARRES2, with shRNA reduced the antibacterial activity of RPE cells. Senescent RPE cells had lower levels of α-secretase and higher levels of β-secretase (BACE1) and γ-secretase (PS1), accompanied by reduced soluble APPα and increased amyloid beta (Aβ) production, particularly the Aβ42 isoform. Eyes from aged donors showed a higher Aβ accumulation within RPE cells. Our results suggest that while RPE cells possess antimicrobial activity, this ability declines with age and is impaired in senescent cells. The impaired antimicrobial activity and augmented Aβ deposition in senescent RPE cells may contribute to age-related retinal para-inflammation and neurodegeneration.