Prolonged hypernutrition impairs TREM2-dependent efferocytosis to license chronic liver inflammation and NASH development.

Obesity-induced chronic liver inflammation is a hallmark of nonalcoholic steatohepatitis (NASH)-an aggressive form of nonalcoholic fatty liver disease. However, it remains unclear how such a low-grade, yet persistent, inflammation is sustained in the liver. Here, we show that the macrophage phagocytic receptor TREM2, induced by hepatocyte-derived sphingosine-1-phosphate, was required for efferocytosis of lipid-laden apoptotic hepatocytes and thereby maintained liver immune homeostasis. However, prolonged hypernutrition led to the production of proinflammatory cytokines TNF and IL-1ß in the liver to induce TREM2 shedding through ADAM17-dependent proteolytic cleavage. Loss of TREM2 resulted in aberrant accumulation of dying hepatocytes, thereby further augmenting proinflammatory cytokine production. This ultimately precipitated a vicious cycle that licensed chronic inflammation to drive simple steatosis transition to NASH. Therefore, impaired macrophage efferocytosis is a previously unrecognized key pathogenic event that enables chronic liver inflammation in obesity. Blocking TREM2 cleavage to restore efferocytosis may represent an effective strategy to treat NASH.

Autophagy, Inflammation, and Immunity: A Troika Governing Cancer and Its Treatment.

Autophagy, a cellular waste disposal process, has well-established tumor-suppressive properties. New studies indicate that, in addition to its cell-autonomous anti-tumorigenic functions, autophagy inhibits cancer development by orchestrating inflammation and immunity. While attenuating tumor-promoting inflammation, autophagy enhances the processing and presentation of tumor antigens and thereby stimulates anti-tumor immunity. Although cancer cells can escape immunosurveillance by tuning down autophagy, certain chemotherapeutic agents with immunogenic properties may enhance anti-tumor immunity by inducing autophagic cell death. Understanding the intricate and complex relationships within this troika and how they are affected by autophagy enhancing drugs should improve the efficacy of cancer immunotherapy.

NF-κB Restricts Inflammasome Activation via Elimination of Damaged Mitochondria.

Nuclear factor κB (NF-κB), a key activator of inflammation, primes the NLRP3-inflammasome for activation by inducing pro-IL-1ß and NLRP3 expression. NF-κB, however, also prevents excessive inflammation and restrains NLRP3-inflammasome activation through a poorly defined mechanism. We now show that NF-κB exerts its anti-inflammatory activity by inducing delayed accumulation of the autophagy receptor p62/SQSTM1. External NLRP3-activating stimuli trigger a form of mitochondrial (mt) damage that is caspase-1- and NLRP3-independent and causes release of direct NLRP3-inflammasome activators, including mtDNA and mtROS. Damaged mitochondria undergo Parkin-dependent ubiquitin conjugation and are specifically recognized by p62, which induces their mitophagic clearance. Macrophage-specific p62 ablation causes pronounced accumulation of damaged mitochondria and excessive IL-1ß-dependent inflammation, enhancing macrophage death. Therefore, the "NF-κB-p62-mitophagy" pathway is a macrophage-intrinsic regulatory loop through which NF-κB restrains its own inflammation-promoting activity and orchestrates a self-limiting host response that maintains homeostasis and favors tissue repair.

Limiting inflammation-the negative regulation of NF-κB and the NLRP3 inflammasome.

A properly mounted immune response is indispensable for recognizing and eliminating danger arising from foreign invaders and tissue trauma. However, the 'inflammatory fire' kindled by the host response must be tightly controlled to prevent it from spreading and causing irreparable damage. Accordingly, acute inflammation is self-limiting and is normally attenuated after elimination of noxious stimuli, restoration of homeostasis and initiation of tissue repair. However, unresolved inflammation may lead to the development of chronic autoimmune and degenerative diseases and cancer. Here, we discuss the key molecular mechanisms that contribute to the self-limiting nature of inflammatory signaling, with emphasis on the negative regulation of the NF-κB pathway and the NLRP3 inflammasome. Understanding these negative regulatory mechanisms should facilitate the development of much-needed therapeutic strategies for treatment of inflammatory and autoimmune pathologies.

The molecular anatomy of cashmere goat hair follicle during cytodifferentiation stage.

BACKGROUND: Cashmere, named as "soft gold", derives from the secondary hair follicles (SHFs) of cashmere goat which is vital to Northwest China's economy. The cytodifferentiation stage (E120), mirroring the complete hair follicle (HF) structure of adult goats and marking a critical phase in SHF development. Therefore, this study aims to enhance the understanding of SHF development and its impact on fiber quality, informing breeding strategies. RESULTS: From the scRNA-seq data analysis, the intricate processes and transcriptional dynamics of inner layer cell differentiation of HFs were unveiled in this study. we identified nine cell populations during cytodifferentiation and key structures such as the hair shaft and inner root sheath. And we discovered three main inner layer lineages and seven subpopulations, clarifying their roles in specialization and signaling. Pseudotime mapping analysis showed cell evolution from early stage to mature stages marked by unique gene expressions, and the intermediate stage on the differentiation of each lineage was revealed. The identification and spatial localization of specific transcription factors, such as GATA3, LEF1 and PRDM1, as well as keratin genes highlight regulatory pathways involved in HF development, which was further validated by immunofluorescence. These findings suggested the potential strategies to improve fiber quality, and the discovery of diverse cell types and their developmental molecular mechanisms, particularly in this species-specific context, offered a nuanced view of the regulatory mechanisms driving HF development in cashmere goats. CONCLUSION: Overall, these findings provide a systematic molecular atlas of skin, defining three major branches and cell states of inner layer cells of HF, and determining how the branch-specific transcription factors, keratins, and signals coordinate HF morphogenesis during cytodifferentiation stage. This research not only advances skin tissue research in goats but also holds broader implications for the understanding of HF regeneration and development across various species.

New mitochondrial DNA synthesis enables NLRP3 inflammasome activation.

Dysregulated NLRP3 inflammasome activity results in uncontrolled inflammation, which underlies many chronic diseases. Although mitochondrial damage is needed for the assembly and activation of the NLRP3 inflammasome, it is unclear how macrophages are able to respond to structurally diverse inflammasome-activating stimuli. Here we show that the synthesis of mitochondrial DNA (mtDNA), induced after the engagement of Toll-like receptors, is crucial for NLRP3 signalling. Toll-like receptors signal via the MyD88 and TRIF adaptors to trigger IRF1-dependent transcription of CMPK2, a rate-limiting enzyme that supplies deoxyribonucleotides for mtDNA synthesis. CMPK2-dependent mtDNA synthesis is necessary for the production of oxidized mtDNA fragments after exposure to NLRP3 activators. Cytosolic oxidized mtDNA associates with the NLRP3 inflammasome complex and is required for its activation. The dependence on CMPK2 catalytic activity provides opportunities for more effective control of NLRP3 inflammasome-associated diseases.

Author Correction: Inflammation-induced IgA+ cells dismantle anti-liver cancer immunity.

In this Article, the sentence: "After 7 months of HFD, MUP-uPA mice developed HCC15, which contained numerous (usually 50-100 per tumour) non-recurrent coding mutations in pathways that are mutated in human HCC (Fig. 2d and Extended Data Fig. 6a).", should have read: "After 7 months of HFD, MUP-uPA mice developed HCC15, which contained numerous (usually 50-100 per tumour) non-recurrent mutations in pathways that are mutated in human HCC (Fig. 2d and Extended Data Fig. 6a).". This has been corrected online. In Extended Data Fig. 6a and b, which show the number of point mutations identified per sample and the mutational signatures, all sequence variants (including non-coding mutations) are shown. Fig. 2d also presents all variants compared to human mutations. In the Supplementary Information to this Amendment, we now provide the comparisons of all variants and coding variants to human mutations.

Emergence of genetic diversity and multi-drug resistant Clostridium perfringens from wild birds.

BACKGROUND: Clostridium perfringens (C. perfringens) is an important zoonotic microorganism that can cause animal and human infections, however information about the prevalence status in wild birds of this pathogenic bacterium is currently limited. RESULT: In this study, 57 strains of C. perfringens were isolated from 328 fecal samples of wild birds. All the isolates were identified as type A and 70.18% of the isolates carried the cpb2 gene. Antimicrobial susceptibility testing showed that and 22.80% of the isolates were classified as multidrug-resistant strains. The MLST analysis of the 57 isolates from wild birds was categorized into 55 different sequence types (STs) and clustered into eight clonal complexes (CCs) with an average of 20.1 alleles and the Simpson Diversity index (Ds) of 0.9812, and revealed a high level of genetic diversity within the C. perfringens populations. Interestingly, the isolates from swan goose were clustered in the same CC while isolates from other bird species were more scattered suggesting that a potential difference in genetic diversity among the C. perfringens populations associated with different bird species. CONCLUSION: C. perfringens exhibits a wide range of host adaptations, varying degrees of antimicrobial resistance, and a high degree of genetic diversity in wild birds. Understanding the prevalence, toxin type, antimicrobial resistance, and genetic diversity of C. perfringens in wildlife populations is essential for developing effective strategies for disease control and management.

Soluble α-synuclein-antibody complexes activate the NLRP3 inflammasome in hiPSC-derived microglia.

Parkinson's disease is characterized by accumulation of α-synuclein (αSyn). Release of oligomeric/fibrillar αSyn from damaged neurons may potentiate neuronal death in part via microglial activation. Heretofore, it remained unknown if oligomeric/fibrillar αSyn could activate the nucleotide-binding oligomerization domain (NOD)-like receptor (NLR) family pyrin domain-containing 3 (NLRP3) inflammasome in human microglia and whether anti-αSyn antibodies could prevent this effect. Here, we show that αSyn activates the NLRP3 inflammasome in human induced pluripotent stem cell (hiPSC)-derived microglia (hiMG) via dual stimulation involving Toll-like receptor 2 (TLR2) engagement and mitochondrial damage. In vitro, hiMG can be activated by mutant (A53T) αSyn secreted from hiPSC-derived A9-dopaminergic neurons. Surprisingly, αSyn-antibody complexes enhanced rather than suppressed inflammasome-mediated interleukin-1ß (IL-1ß) secretion, indicating these complexes are neuroinflammatory in a human context. A further increase in inflammation was observed with addition of oligomerized amyloid-ß peptide (Aß) and its cognate antibody. In vivo, engraftment of hiMG with αSyn in humanized mouse brain resulted in caspase-1 activation and neurotoxicity, which was exacerbated by αSyn antibody. These findings may have important implications for antibody therapies aimed at depleting misfolded/aggregated proteins from the human brain, as they may paradoxically trigger inflammation in human microglia.

Improved REBA: deep learning based rapid entire body risk assessment for prevention of musculoskeletal disorders.

Preventing work-related musculoskeletal disorders (WMSDs) is crucial in reducing their impact on individuals and society. However, the existing mainstream 2D image-based approach is insufficient in capturing the complex 3D movements and postures involved in many occupational tasks. To address this, an improved deep learning-based rapid entire body assessment (REBA) method has been proposed. The method takes working videos as input and automatically outputs the corresponding REBA score through 3D pose reconstruction. The proposed method achieves an average precision of 94.7% on real-world data, which is comparable to that of ergonomic experts. Furthermore, the method has the potential to be applied across a wide range of industries as it has demonstrated good generalisation in multiple scenarios. The proposed method offers a promising solution for automated and accurate risk assessment of WMSDs, with implications for various industries to ensure the safety and well-being of workers.

This paper proposes a deep learning-based improved rapid entire body assessment (REBA) method for assessing work-related musculoskeletal disorders (WMSDs) risks using 3D pose reconstruction from videos, achieving 94.7% precision, comparable to ergonomic experts, with potential applications across various industries.

Identification of FCER1G as a cyclosporin A plus corticosteroid sensitization gene in female patients with Vogt-Koyanagi-Harada disease.

The resistance development of the combination regimen of corticosteroids (CS) with cyclosporin A (CsA) leads to therapeutic failure of some patients with autoimmune diseases. In the male patients with Vogt-Koyanagi-Harada (VKH) disease, we have identified RPS4Y1 as an important resistance gene of the regimen and a functional mediator of chlorambucil (CLB). However, it remains unclear what is responsible for the resistance in female patients. In the present study, we performed RNA sequencing, tandem mass tag (TMT) proteomics, gain- and loss-of-function assays and rescue assays to screen and validate potential resistant mediators. The results showed that only Fc epsilon receptor Ig (FCER1G) exhibited significantly differential expression in CD4+ T cells among female CsA & CS resistant, sensitive and CLB & CsA & CS treated patients at transcription and protein levels. Inhibition of FCER1G was demonstrated to modulate CD4+ T cell resistance to CsA & CS in female patients. Importantly, the inhibition was mediated by elevated DNA methylation in the promoter region of the FCER1G gene. Moreover, we found that the salvage effect of CLB on CsA & CS resistance was mediated by an increased FCER1G expression via DNA demethylation in female patients. Taken together, the downregulation of FCER1G due to DNA hypermethylation is responsible for the resistance to CsA & CS and CLB reverses this resistance by inducing FCER1G expression via DNA demethylation in female patients. Modulation of FCER1G would be a promising sensitization strategy in female patients with resistance to CsA & CS.

Molecular diagnostic yield for Blau syndrome in previously diagnosed juvenile idiopathic arthritis with uveitis or cutaneous lesions.

OBJECTIVE: Diagnostic pitfalls often arise in the community because of potentially misleading similarities between juvenile idiopathic arthritis (JIA) and Blau syndrome, an immune-related disorder caused by NOD2 gene mutations. It remains unclear in which population and to which extent next-generation sequencing techniques can aid in diagnosis. METHODS: We evaluated clinical usefulness of targeted next-generation sequencing in previously diagnosed JIA. Participants were required to have symptoms and signs suspected of Blau syndrome, including at least uveitis or cutaneous lesions in addition to arthritis. Targeted sequencing was conducted on NOD2 gene to detect diagnostic variants classified as pathogenic or likely pathogenic for Blau syndrome. We assessed the molecular diagnostic yield and clinical implications on patient care. RESULTS: Between May 1, 2008, and June 1, 2021, sequencing data were accrued from 123 previously diagnosed JIA (median age: 5 years; female: 62.6%). Targeted NOD2 sequencing yielded a positive molecular diagnosis of Blau syndrome in 21.1% (95% CI, 14.9%-29.2%), encompassing six heterozygous missense mutations classified as pathogenic variants. Among those receiving a molecular diagnosis, changes in clinical management and treatment were considered as having occurred in 38.5%. Nine predictors were identified to be associated with a higher diagnostic yield, providing clinical clues to suspect the possibility of Blau syndrome. CONCLUSION: Among some patients with pediatric-onset arthritis complicated with uveitis or cutaneous lesions, reassessing their diagnosis of JIA may be warranted. Targeted NOD2 sequencing established the molecular diagnosis of Blau syndrome in nearly one fifth of these cases and provided clinically relevant information for patient-care decisions.

Interleukin-17 receptor a signaling in transformed enterocytes promotes early colorectal tumorigenesis.

Interleukin-17A (IL-17A) is a pro-inflammatory cytokine linked to rapid malignant progression of colorectal cancer (CRC) and therapy resistance. IL-17A exerts its pro-tumorigenic activity through its type A receptor (IL-17RA). However, IL-17RA is expressed in many cell types, including hematopoietic, fibroblastoid, and epithelial cells, in the tumor microenvironment, and how IL-17RA engagement promotes colonic tumorigenesis is unknown. Here we show that IL-17RA signals directly within transformed colonic epithelial cells (enterocytes) to promote early tumor development. IL-17RA engagement activates ERK, p38 MAPK, and NF-κB signaling and promotes the proliferation of tumorigenic enterocytes that just lost expression of the APC tumor suppressor. Although IL-17RA signaling also controls the production of IL-6, this mechanism makes only a partial contribution to colonic tumorigenesis. Combined treatment with chemotherapy, which induces IL-17A expression, and an IL-17A neutralizing antibody enhanced the therapeutic responsiveness of established colon tumors. These findings establish IL-17A and IL-17RA as therapeutic targets in colorectal cancer.

Inflammation-induced IgA+ cells dismantle anti-liver cancer immunity.

The role of adaptive immunity in early cancer development is controversial. Here we show that chronic inflammation and fibrosis in humans and mice with non-alcoholic fatty liver disease is accompanied by accumulation of liver-resident immunoglobulin-A-producing (IgA+) cells. These cells also express programmed death ligand 1 (PD-L1) and interleukin-10, and directly suppress liver cytotoxic CD8+ T lymphocytes, which prevent emergence of hepatocellular carcinoma and express a limited repertoire of T-cell receptors against tumour-associated antigens. Whereas CD8+ T-cell ablation accelerates hepatocellular carcinoma, genetic or pharmacological interference with IgA+ cell generation attenuates liver carcinogenesis and induces cytotoxic T-lymphocyte-mediated regression of established hepatocellular carcinoma. These findings establish the importance of inflammation-induced suppression of cytotoxic CD8+ T-lymphocyte activation as a tumour-promoting mechanism.

Erratum: Inflammation-induced IgA+ cells dismantle anti-liver cancer immunity.

This corrects the article DOI: 10.1038/nature24302.

Vogt-Koyanagi-Harada disease in pediatric, adult and elderly: clinical characteristics and visual outcomes.

PURPOSE: To depict a whole spectrum of clinical feartures and visual prognosis among pediatric, adult, and elderly Vogt-Koyanagi-Harada disease (VKH) patients. METHODS: Retrospective chart review was conducted in 2571 VKH patients diagnosed from April 2008 to January 2022. Based on age of disease onset, patients were divided into pediatric (age ≤ 16 years), adult (16 < age < 65 years), and elderly (age ≥ 65 years) VKH group. Ocular and extraocular manifestations were compared among these patients. Visual outcomes and complications were evaluated using logistic regression models and restricted cubic splines analysis. RESULTS: The median follow-up time was 48 (IQR, 12-60) months. Pediatric, adult and elderly VKH were found in 106 (4.1%), 2355 (91.6%), and 110 (4.3%) patients, respectively. All of the patients showed similar ocular manifestations in the context of disease phasing. The proportion of neurological and auditory manifestations in pediatric (42.3% and 7.5%) VKH patients was significantly lower than that in adults (66.5% and 47.9%) and elderly (68.2% and 50%) (both p < 0.0001). An increased risk of macular abnormalities was seen in adults (OR, 3.43; 95% CI, 1.62-7.29) compared with elderly VKH. An inverted-U-shaped pattern was observed between disease onset age and a poor visual outcome (visual acuity 6/18 or worse) according to OR value in VKH patients. The highest risk of BCVA ≤ 6/18 was observed in 32 years at disease onset (OR, 1.51; 95% CI, 1.18-1.94). A higher risk of visual loss was observed in adult VKH patients (OR, 9.06; 95% CI, 2.18-37.6) compared with elderly VKH patients. And stratified by macular abnormalities, the interaction test was not significant (P = 0.634). CONCLUSION: Our study identified, for the first time, a whole spectrum of clinical features of VKH based on a large cohort of Chinese patients. Adult VKH patients have an increased risk of poor visual outcomes, possibly due to increased frequency of macular abnormalities.

Average corticosteroid dose and risk for HBV reactivation and hepatitis flare in patients with resolved hepatitis B infection.

OBJECTIVES: Corticosteroids remain the mainstay of treatment for rheumatic diseases but can cause hepatitis B virus (HBV) reactivation in patients with resolved HBV infection. Risk assessment and stratification are needed to guide the management of these patients before corticosteroid therapy. METHODS: We prospectively enrolled patients with negative hepatitis B surface antigen positive Anti-hepatitis B core status with or without corticosteroid use and determined corticosteroid exposure by calculating cumulative dose and time-weighted average daily dose of prednisone. The primary outcome was the time to a composite of HBV reactivation, hepatitis flare or severe hepatitis. RESULTS: Among 1303 participants, the median of cumulative dose and time-weighted average dose of prednisone used in this cohort was 3000 mg (IQR: 300-6750 mg) and 15 mg/day (IQR: 10-20 mg/day), respectively. In multivariable analyses, cumulative dose showed inverted V-shaped relationship with primary events, which peaked at a cumulative dose of 1506 mg (HR: 3.72; 95% CI, 1.96 to 7.08). Quartiles of time-weighted average dose were independently associated with a monotonic increase in event risk (HR per quartile increase: 2.15; 95% CI, 1.56 to 2.98), reaching an HR of 49.48 (95% CI, 6.24 to 392.48) in the top quartile. The incidence of primary outcome was 16.67 per 100 person-years in the top quartile of time-weighted average dose (Q4>20 mg/day). Other quartiles all had an incidence of primary outcome less than 10 per 100 person-years. CONCLUSION: Patients with time-weighted average prednisone dose greater than 20 mg/day would be classified as the high risk for HBV reactivation or hepatitis flare. Prophylactic Anti-HBV therapy may be needed for these high-risk patients. TRIAL REGISTRATION NUMBER: ChiCTR1900023955.

Risk for uveitis relapse after COVID-19 vaccination.

OBJECTIVES: Several studies suggested that coronavirus disease 2019 (COVID-19) vaccination may lead to uveitis, a vision-threatening condition often associated with a variety of autoimmune or autoinflammatory diseases. This study aims to explore factors that influence the risk of uveitis relapse after COVID-19 vaccination to guide the prevention of disease. METHODS: Uveitis relapse was evidenced by worsening activity of intraocular inflammation (e.g. anterior chamber cells, vitreous haze) as defined by the Standardization of Uveitis Nomenclature Working Group. Time to uveitis relapse since the administration of each dose of COVID-19 vaccine was compared across participants with modifiable variables. RESULTS: The primary analysis included 438 non-COVID-19 participants with 857 doses of COVID-19 vaccine administered in total. The median age was 41 years (interquartile range, 30 to 51), and 57.3% were female. A total of 39 episodes of uveitis relapse events occurred in 34 patients after the receipt of a dose of COVID-19 vaccine within 30 days. The median time to relapse after vaccination was 5 days (interquartile range, 1 to 14). Concomitant use of systemic glucocorticoids at the time of vaccination was independently associated with a decrease in risk of relapse after vaccination (HR, 0.23 [95% CI, 0.07-0.74]; P value = 0.014). There was a trend in attenuating the risk of relapse with increasing prednisone dose from none to less than 20 mg per day and then to 20 mg per day or greater (P value for trend = 0.029). CONCLUSIONS: Concomitant treatment with systemic glucocorticoids for uveitis at the time of COVID-19 vaccination was associated with a dose-dependent lower risk of uveitis relapse after vaccination.

Evaluation of sensitivity and specificity of diagnostic criteria for Behçet's disease in the absence of a gold standard.

OBJECTIVE: The performance of existing diagnostic criteria for Behçet's disease (BD) is usually evaluated by comparison with expert opinions, which may be limited by misclassification and disagreement among experts. We aim to evaluate these criteria in the absence of a gold standard. METHODS: We obtained two datasets involving possible BD and other mimickers from a uveitis registry using case-cohort and nested case-control analyses, respectively. With a Bayesian inference approach, the sensitivity and specificity of International Study Group (ISG) and International Criteria for Behçet's Disease (ICBD) criteria were simultaneously estimated when true BD state was unknown. RESULTS: A total of 2440 and 2224 participants were included in case-cohort and nested case-control analyses, respectively. In case-cohort analysis, with scores of ≥4 for BD diagnosis, ICBD criteria showed higher sensitivity (median 97.6%; 95% credible interval 96.9, 98.2) than ISG criteria (median 90.0%; 95% credible interval 88.8, 91.2) but had lower specificity (median 90.8%; 95% credible interval 89.4, 92.1) than ISG criteria (median 98.8%; 95% credible interval 98.3, 99.3). With scores of ≥5 for diagnosis, ICBD criteria demonstrated higher sensitivity (median 97.5%; 95% credible interval 96.8, 98.1) and specificity (median 99.6%; 95% credible interval 99.3, 99.8) than the sensitivity (median 92.3%; 95% credible interval 91.2, 93.3) and specificity (median 98.8%; 95% credible interval 98.2, 99.2) for ISG criteria. The highest diagnostic consistency was observed between ISG criteria and ICBD criteria with scores of ≥5 for diagnosis (Kappa = 0.999; P < 0.001). Nested case-control analysis showed similar results. CONCLUSION: ICBD criteria showed optimum discriminatory properties in sensitivity and specificity with scores of ≥5 for BD diagnosis in uveitis. The diagnostic threshold of ICBD criteria could be considered adjustable according to medical specialty, disease prevalence and local practice characteristics.

Genetic and Clinical Features of Blau Syndrome among Chinese Patients with Uveitis.

PURPOSE: The American College of Medical Genetics and Genomics (ACMG) and the Association for Molecular Pathology call for cautious interpretation of variants as causative of a monogenic disorder by stringent standards. We aimed to reclassify the pathogenicity of nucleotide binding oligomerization domain containing 2 (NOD2) variants according to the ACMG guidelines and to characterize clinical features in patients whose ocular disease might actually be explained by Blau syndrome. DESIGN: Genetic analysis and descriptive study. PARTICIPANTS: A total of 1003 unrelated healthy individuals and 3921 sporadic patients who presented with uveitis. METHODS: Whole-exome sequencing was performed on all healthy participants and 551 patients with uveitis, and targeted NOD2 resequencing was performed on the remaining 3370 patients with uveitis. Pathogenicity for Blau syndrome was classified for NOD2 variants identified by sequencing in study participants according to the ACMG guidelines. Clinical manifestations were compared among NOD2 variants of different levels of classification. MAIN OUTCOME MEASURES: Pathogenicity of variants. RESULTS: Eight NOD2 gain-of-function mutations, p.R334W, p.R334Q, p.E383K, p.G481D, p.W490S, p.M513T, p.R587C, and p.N670K, were classified as pathogenic, and 66 patients (1.7%) with uveitis were diagnosed with Blau syndrome due to these mutations. Of 66 with Blau syndrome, anterior uveitis accounted for 39.4%, posterior uveitis for 9.1%, and panuveitis for 51.5%. A proportion of 21.2% of Blau syndrome presented as multifocal choroiditis, 48.5% had papillitis, and 74.2% showed retinal microvasculitis detected by fundus fluorescein angiography. Six NOD2 variants, p.P268S, p.R311W, p.R471C, p.A612T, p.R702W, and p.V955I, were considered nonpathogenic for Blau syndrome and were identified in 96 patients with uveitis. The incidence of bilateral uveitis (86.4%), secondary glaucoma (47.0%), epiretinal membrane (7.6%), choroidal neovascularization (4.6%), retinal atrophy (10.6%), arthritis (69.7%), joint deformity (51.5%), and skin rash (40.9%) was higher in Blau syndrome than in patients with uveitis carrying non-Blau-causing NOD2 variants. Patients with Blau syndrome permanently experienced overall poorer best-corrected visual acuity. Several rare NOD2 mutations, p.I722L (2 cases), p.T476P (1 case), p.T476del (1 case), and p.R439H (1 case), were newly identified. CONCLUSIONS: Pathogenic NOD2 variants for Blau syndrome were limited to those gain-of-function mutations and were associated with a high risk for arthritis, skin rash, permanent visual loss, and ocular complications in patients with uveitis.