TNFAIP3 Overexpression Inhibits Diffuse Large B-Cell Lymphoma Progression by Promoting Autophagy through TLR4/MyD88/NF-κB Signaling Pathway.

BACKGROUND: Tumor necrosis factor alpha induced protein 3 (TNFAIP3) is reportedly to have significant implications for autophagy regulation in various cancers. The current study aimed to decipher the role and mechanism of TNFAIP3 in diffuse large B-cell lymphoma (DLBCL) by modulating autophagy. METHODS: Information pertaining to the differential expression and prognostic role of TNFAIP3 in DLBCL was gleaned from the Gene Expression Omnibus (GEO) database. The TNFAIP3 expression levels in human DLBCL cells were detected by quantitative real-time polymerase chain reaction (qRT-PCR) and Western blotting. Cell counting kit-8 (CCK-8) and colony formation assays were employed to determine cell proliferation. Transwell assay and flow cytometry were applied to detect cell migration and apoptosis, respectively. Immunofluorescence and transmission electron microscope were used for the assessment of cell autophagy. The levels of apoptotic markers (caspase-3, cleaved-caspase-3, Bcl-2 Associated X (Bax), and B cell lymphoma-2 (Bcl-2)), autophagy indicators (the ratio of microtubule-associated proteins 1A/1B light chain 3 II and I (LC3II/LC3I), Sequestosome (p62)), and pathway proteins (toll-like receptor 4 (TLR4), myeloid differentiation primary response 88 (MyD88), Transcription Factor NF-Kappa-B P65 Subunit (p65), and phosphorylated-p65 (p-p65)) were assessed via Western blotting. Immunohistochemistry was employed to detect Ki67 expression in tumor tissues. RESULTS: TNFAIP3 expression in DLBCL samples was downregulated, correlating with poor prognosis. TNFAIP3 expression was also downregulated in DLBCL cells. It was found that TNFAIP3 impeded cell proliferation and migration, and enhanced apoptosis of OCI-LY3 cells. Intervention with autophagy inhibitor 3-methyladenine (3-MA) markedly reversed apoptosis of OCI-LY3 cells induced by TNFAIP3. Besides, TNFAIP3 induced autophagy via modulating the TLR4/MyD88/nuclear factor kappa B (NF-κB) signaling pathway. In vivo experiments showed that TNFAIP3 expression in DLBCL was downregulated, and upregulation of TNFAIP3 could inhibit tumor growth. CONCLUSION: TNFAIP3 inhibits DLBCL progression by inducing TLR4/MyD88/NF-κB pathway-mediated autophagy.

Genetic Mutations Associated With TNFAIP3 (A20) Haploinsufficiency and Their Impact on Inflammatory Diseases.

TNF-α-induced protein 3 (TNFAIP3), commonly referred to as A20, is an integral part of the ubiquitin-editing complex that significantly influences immune regulation, apoptosis, and the initiation of diverse immune responses. The A20 protein is characterized by an N-terminal ovarian tumor (OTU) domain and a series of seven zinc finger (ZNF) domains. Mutations in the TNFAIP3 gene are implicated in various immune-related diseases, such as Behçet's disease, polyarticular juvenile idiopathic arthritis, autoimmune thyroiditis, autoimmune hepatitis, and rheumatoid arthritis. These mutations can lead to a spectrum of symptoms, including, but not limited to, recurrent fever, ulcers, rashes, musculoskeletal and gastrointestinal dysfunctions, cardiovascular issues, and respiratory infections. The majority of these mutations are either nonsense (STOP codon) or frameshift mutations, which are typically associated with immune dysfunctions. Nonetheless, missense mutations have also been identified as contributors to these conditions. These genetic alterations may interfere with several biological pathways, notably abnormal NF-κB signaling and dysregulated ubiquitination. Currently, there is no definitive treatment for A20 haploinsufficiency; however, therapeutic strategies can alleviate the symptoms in patients. This review delves into the mutations reported in the TNFAIP3 gene, the clinical progression in affected individuals, potential disease mechanisms, and a brief overview of the available pharmacological interventions for A20 haploinsufficiency. Mandatory genetic testing of the TNFAIP3 gene should be performed in patients diagnosed with autoinflammatory disorders to better understand the genetic underpinnings and guide treatment decisions.

A20 in hepatic stellate cells suppresses chronic hepatitis by inhibiting DCLK1-JNK pathway-dependent chemokines.

Hepatic stellate cells (HSCs) are responsible for liver fibrosis accompanied by its activation into myofibroblasts and the abundant production of extracellular matrix. However, the HSC contribution to progression of liver inflammation has been less known. We aimed to elucidate the mechanism in HSCs underlying the inflammatory response and the function of tumor necrosis factor α-related protein A20 (TNFAIP3). We established A20 conditional knockout (KO) mice crossing Twist2-Cre and A20 floxed mice. Using these mice, the effect of A20 was analyzed in mouse liver and HSCs. The human HSC line LX-2 was also used to examine the role and underlying molecular mechanism of A20. In this KO model, A20 was deficient in >80% of HSCs. Spontaneous inflammation with mild fibrosis was found in the liver of the mouse model without any exogenous agents, suggesting that A20 in HSCs suppresses chronic hepatitis. Comprehensive RNA sequence analysis revealed that A20-deficient HSCs exhibited an inflammatory phenotype and abnormally expressed chemokines. A20 suppressed JNK pathway activation in HSCs. Loss of A20 function in LX-2 cells also induced excessive chemokine expression, mimicking A20-deficient HSCs. A20 overexpression suppressed chemokine expression in LX-2. In addition, we identified DCLK1 in the genes regulated by A20. DCLK1 activated the JNK pathway and upregulates chemokine expression. DCLK1 inhibition significantly decreased chemokine induction by A20-silencing, suggesting that A20 controlled chemokine expression in HSCs via the DCLK1-JNK pathway. In conclusion, A20 suppresses chemokine induction dependent on the DCLK1-JNK signaling pathway. These findings demonstrate the therapeutic potential of A20 and the DCLK1-JNK pathway for the regulation of inflammation in chronic hepatitis.

Structural basis for TNIP1 binding to FIP200 during mitophagy.

TNIP1 has been increasingly recognized as a security check to finely adjust the rate of mitophagy by disrupting the recycling of the Unc-51-like kinase complex during autophagosome formation. Through tank-binding kinase 1-mediated phosphorylation of the TNIP1 FIP200 interacting region (FIR) motif, the binding affinity of TNIP1 for FIP200, a component of the Unc-51-like kinase complex, is enhanced, allowing TNIP1 to outcompete autophagy receptors. Consequently, FIP200 is released from the autophagosome, facilitating further autophagosome expansion. However, the molecular basis by which FIP200 utilizes its claw domain to distinguish the phosphorylation status of residues in the TNIP1 FIR motif for recognition is not well understood. Here, we elucidated multiple crystal structures of the complex formed by the FIP200 claw domain and various phosphorylated TNIP1 FIR peptides. Structural and isothermal titration calorimetry analyses identified the crucial residues in the FIP200 claw domain responsible for the specific recognition of phosphorylated TNIP1 FIR peptides. Additionally, utilizing structural comparison and molecular dynamics simulation data, we demonstrated that the C-terminal tail of TNIP1 peptide affected its binding to the FIP200 claw domain. Moreover, the phosphorylation of TNIP1 Ser123 enabled the peptide to effectively compete with the peptide p-CCPG1 (the FIR motif of the autophagy receptor CCPG1) for binding with the FIP200 claw domain. Overall, our work provides a comprehensive understanding of the specific recognition of phosphorylated TNIP1 by the FIP200 claw domain, marking an initial step toward fully understanding the molecular mechanism underlying the TNIP1-dependent inhibition of mitophagy.

Molecular profiling of core immune-escape genes highlights TNFAIP3 as an immune-related prognostic biomarker in neuroblastoma.

BACKGROUND: Neuroblastoma (NB) is the most prevalent and deadliest pediatric solid tumor. With of over 50% of high-risk neuroblastoma cases relapse, the imperative for novel drug targets and therapeutic strategies is accentuated. In neuroblastoma, the existence of tumor-associated macrophages (TAMs) correlates with an unfavorable patient prognosis. However, the clinical relevance and prognostic implications of regulatory genes linked to TAMs infiltration in neuroblastoma remain unclear, and further study is required. METHODS: We conducted a comprehensive analysis utilizing transcriptome expression profiles from three primary datasets associated with neuroblastoma (GSE45547, GSE49710, TARGET) to identify hub genes implicated in immune evasion within neuroblastoma. Subsequently, we utilized single-cell RNA sequencing analysis on 17 clinical neuroblastoma samples to investigate the expression and distribution of these hub genes, leading to the identification of TNFAIP3. The above three public databases were merged to allowed for the validation of TNFAIP3's molecular functions through GO and KEGG analysis. Furthermore, we assessed TNFAIP3's correlation with immune infiltration and its potential immunotherapeutic impact by multiple algorithms. Our single-cell transcriptome data revealed the role of TNFAIP3 in macrophage polarization. Finally, preliminary experimental verifications to confirm the biological functions of TNFAIP3-mediated TAMs in NB. RESULTS: A total of 6 genes related to immune evasion were screened and we found that TNFAIP3 exhibited notably higher expression in macrophages than other immune cell types, based on the scRNA-sequencing data. GO and KEGG analysis showed that low expression of TNFAIP3 significantly correlated with the activation of multiple oncogenic pathways as well as immune-related pathways. Then validation affirmed that individuals within the TNFAIP3 high-expression cohort could potentially derive greater advantages from immunotherapeutic interventions, alongside exhibiting heightened immune responsiveness. Deciphering the pseudotime trajectory of macrophages, we revealed the potential of TNFAIP3 in inducing the polarization of macrophages towards the M1 phenotype. Finally, we confirmed that patients in the TNFAIP3 high expression group might benefit more from immunotherapy or chemotherapy as substantiated by RT-qPCR and immunofluorescence examinations. Moreover, the role of TNFAIP3 in macrophage polarization was validated. Preliminary experiment showed that TNFAIP3-mediated TAMs inhibit the proliferation, migration and invasion capabilities of NB cells. CONCLUSIONS: Our results suggest that TNFAIP3 was first identified as a promising biomarker for immunotherapy and potential molecular target in NB. Besides, the presence of TNFAIP3 within TAMs may offer a novel therapeutic strategy for NB.

A20 in Kidney Transplantation and Autoimmunity.

A20, the central inhibitor of NFκB, has multiple anti-inflammatory properties, making it an interesting target in kidney autoimmune disease and transplant biology. It has been shown to be able to inhibit inflammatory functions in macrophages, dendritic cells, T cells, and B cells in various ways, leading to less tissue damage and better graft outcomes. In this review, we will discuss the current literature regarding A20 in kidney transplantation and autoimmunity. Future investigations on animal models and in existing immunosuppressive therapies are needed to establish A20 as a therapeutic target in kidney transplantation and autoimmunity. Cell-based therapies, modified viruses or RNA-based therapies could provide a way for A20 to be utilized as a promising mediator of inflammation and tissue damage.

Inhibition of NLRP3 inflammasome activation by A20 through modulation of NEK7.

The NLRP3 inflammasome, a pivotal component of innate immunity, has been implicated in various inflammatory disorders. The ubiquitin-editing enzyme A20 is well known to regulate inflammation and maintain homeostasis. However, the precise molecular mechanisms by which A20 modulates the NLRP3 inflammasome remain poorly understood. Here, our study revealed that macrophages deficient in A20 exhibit increased protein abundance and elevated mRNA level of NIMA-related kinase 7 (NEK7). Importantly, A20 directly binds with NEK7, mediating its K48-linked ubiquitination, thereby targeting NEK7 for proteasomal degradation. Our results demonstrate that A20 enhances the ubiquitination of NEK7 at K189 and K293 ubiquitinated sites, with K189 playing a crucial role in the binding of NEK7 to A20, albeit not significantly influencing the interaction between NEK7 and NLRP3. Furthermore, A20 disrupts the association of NEK7 with the NLRP3 complex, potentially through the OTU domain and/or synergistic effect of ZnF4 and ZnF7 motifs. Significantly, NEK7 deletion markedly attenuates the activation of the NLRP3 inflammasome in A20-deficient conditions, both in vitro and in vivo. This study uncovers a mechanism by which A20 inhibits the NLRP3 inflammasome.

Dysregulation of TNF-induced protein 3 and CCAAT/enhancer-binding protein ß in alveolar macrophages: Implications for systemic sclerosis-associated interstitial lung disease.

OBJECTIVES: This study investigates the role of TNF-induced protein 3 (TNFAIP3) and CCAAT/enhancer-binding protein ß (C/EBPß) in alveolar macrophages (AMs) of patients with systemic sclerosis-associated interstitial lung disease (SSc-ILD) and their influence on pulmonary fibrosis. METHODS: Transfection of HEK293T cells and AMs with plasmids carrying TNFAIP3 and C/EBPß was performed, followed by co-culturing AMs with pulmonary fibroblasts. Immunoblotting analysis was then utilized to assess the expression of TNFAIP3, C/EBPß, and collagen type 1 (Col1). Quantitative PCR analysis was conducted to quantify the mRNA levels of C/EBPß, IL-10, and TGF-ß1. STRING database analysis, and immunoprecipitation assays were employed to investigate the interactions between TNFAIP3 and C/EBPß. RESULTS: TNFAIP3 expression was significantly reduced in SSc-ILD AMs, correlating with increased Col1 production in fibroblasts. Overexpression of TNFAIP3 inhibited this pro-fibrotic activity. Conversely, C/EBPß expression was elevated in SSc-ILD AMs, and its reduction through TNFAIP3 restoration decreased pro-fibrotic cytokines IL-10 and TGFß1 levels. Protein-protein interaction studies confirmed the regulatory relationship between TNFAIP3 and C/EBPß. CONCLUSIONS: This study highlights the important role of TNFAIP3 in regulating pulmonary fibrosis in SSc-ILD by modulating C/EBPß expression in AMs. These findings suggest that targeting TNFAIP3 could be a potential therapeutic strategy for managing SSc-ILD patients.

Interactions between TNFAIP3, PTPN22, and TRAF1-C5 gene polymorphisms in patients with primary Sjögren's syndrome.

Objectives: The aim of our study was to investigate whether TNFAIP3, PTPN22, and TRAF1-5 single nucleotide polymorphisms (SNPs) are associated with susceptibility, severity, or serological markers in primary Sjögren's syndrome (pSS). Patients and methods: The cases and controls study was conducted between December 2021 and June 2022. TNFAIP3 rs10499194C/T, rs6920220G/A, and rs2230926T/G, PTPN22 rs2476601C/T and rs33996649G/A, and TRAF1-C5 rs10818488G/A polymorphisms were genotyped in 154 female pSS patients (mean age: 45.2±6.8 years) and 313 female control subjects (mean age: 50.3±7.5 years) using the TaqMan® SNP genotyping assay. An association analysis between TNFAIP3, PTPN22, and TRAF1-C5 SNPs and susceptibility, clinical characteristics, and serological markers of pSS was performed. Interactions between TNFAIP3, PTPN22, and TRAF1-C5 SNPs were also evaluated in patients and controls. Results: The genotype and allele frequencies showed no association with susceptibility, severity, or serological markers of pSS. Nevertheless, several interactions between TNFAIP3 and TRAF1-C5 or TNFAIP3, PTPN22, and TRAF1-C5 genotypes were associated with susceptibility to pSS (p<0.01). Conclusion: Individual TNFAIP3, PTPN22, and TRAF1-C5 SNPs are not associated with susceptibility, severity, or serological markers of pSS. However, genetic interactions between TRAF1-C5 and TNFAIP3 or TNFAIP3, PTPN22, and TRAF1-C5 SNPs are risk factors for pSS.

Tumor necrosis factor alpha induced protein 3, interleukin 10, tumor necrosis factor alpha, and interleukin 17 F genes polymorphisms in Algerian patients with rheumatoid arthritis.

BACKGROUND: Rheumatoid arthritis (RA) is a systemic autoimmune disease with chronic inflammation. Its pathogenesis involves immunological, genetic, and environmental factors. We investigate the association between Tumor Necrosis Factor α Protein 3 (TNFAIP3), Interleukin 10 (IL10), Tumor Necrosis Factor α (TNF α), and Interleukin 17 F (IL17F) polymorphisms with susceptibility to RA. METHODS AND RESULTS: 191 patients with RA diagnosed according to the American College of Rheumatology (ACR)/ European League Against Rheumatism (EULAR) classification and 190 healthy subjects were recruited. Rheumatoid factor (RF), anti-citrullinated peptide antibodies (ACPA), and C-reactive protein (CRP) were measured. Genotyping of the polymorphisms was performed by real-time PCR. Analysis of the allelic frequencies of TNFAIP3 showed a positive association OR (95% CI) = 1.46 (1.01-2.09); p = 0.04, but failed to meet the criteria of significance after Bonferroni Correction. The genotypic and allelic distribution of the IL10, IL17F, and TNFα showed no significant difference when comparing the RA group with controls. Furthermore, the genotype codominant model shows a moderate positive association in the presence of ACPA (OR (95% CI) = 2.82 (1.22-6.24); p = 0.01. None of the polymorphisms studied was associated with RF and CRP production. CONCLUSION: Our results show that there is a tendency for the AG genotype of IL10-1082 to be associated with the production of ACPA in patients with RA. None of the variants studied were associated with RA susceptibility in Algerians.

Myeloid A20 is critical for alternative macrophage polarization and type-2 immune-mediated helminth resistance.

Background: Protective immunity against intestinal helminths requires induction of robust type-2 immunity orchestrated by various cellular and soluble effectors which promote goblet cell hyperplasia, mucus production, epithelial proliferation, and smooth muscle contractions to expel worms and re-establish immune homeostasis. Conversely, defects in type-2 immunity result in ineffective helminth clearance, persistent infection, and inflammation. Macrophages are highly plastic cells that acquire an alternatively activated state during helminth infection, but they were previously shown to be dispensable for resistance to Trichuris muris infection. Methods: We use the in vivo mouse model A20myel-KO, characterized by the deletion of the potent anti-inflammatory factor A20 (TNFAIP3) specifically in the myeloid cells, the excessive type-1 cytokine production, and the development of spontaneous arthritis. We infect A20myel-KO mice with the gastrointestinal helminth Trichuris muris and we analyzed the innate and adaptive responses. We performed RNA sequencing on sorted myeloid cells to investigate the role of A20 on macrophage polarization and type-2 immunity. Moreover, we assess in A20myel-KO mice the pharmacological inhibition of type-1 cytokine pathways on helminth clearance and the infection with Salmonella typhimurium. Results: We show that proper macrophage polarization is essential for helminth clearance, and we identify A20 as an essential myeloid factor for the induction of type-2 immune responses against Trichuris muris. A20myel-KO mice are characterized by persistent Trichuris muris infection and intestinal inflammation. Myeloid A20 deficiency induces strong classical macrophage polarization which impedes anti-helminth type-2 immune activation; however, it promotes detrimental Th1/Th17 responses. Antibody-mediated neutralization of the type-1 cytokines IFN-γ, IL-18, and IL-12 prevents myeloid-orchestrated Th1 polarization and re-establishes type-2-mediated protective immunity against T. muris in A20myel-KO mice. In contrast, the strong Th1-biased immunity in A20myel-KO mice offers protection against Salmonella typhimurium infection. Conclusions: We hereby identify A20 as a critical myeloid factor for correct macrophage polarization and appropriate adaptive mucosal immunity in response to helminth and enteric bacterial infection.

The Complexity of Being A20: From Biological Functions to Genetic Associations.

A20, encoded by TNFAIP3, is a critical negative regulator of immune activation. A20 is a ubiquitin editing enzyme with multiple domains, each of which mediates or stabilizes a key ubiquitin modification. A20 targets diverse proteins that are involved in pleiotropic immunologic pathways. The complexity of A20-mediated immunomodulation is illustrated by the varied effects of A20 deletion in different cell types and disease models. Clinically, the importance of A20 is highlighted by its extensive associations with human disease. A20 germline variants are associated with a wide range of inflammatory diseases, while somatic mutations promote development of B cell lymphomas. More recently, the discovery of A20 haploinsufficiency (HA20) has provided real world evidence for the role of A20 in immune cell function. Originally described as an autosomal dominant form of Behcet's disease, HA20 is now considered a complex inborn error of immunity with a broad spectrum of immunologic and clinical phenotypes.

CM1, a Chrysin Derivative, Protects from Endotoxin-Induced Lethal Shock by Regulating the Excessive Activation of Inflammatory Responses.

Sepsis, a leading cause of death worldwide, is a harmful inflammatory condition that is primarily caused by an endotoxin released by Gram-negative bacteria. Effective targeted therapeutic strategies for sepsis are lacking. In this study, using an in vitro and in vivo mouse model, we demonstrated that CM1, a derivative of the natural polyphenol chrysin, exerts an anti-inflammatory effect by inducing the expression of the ubiquitin-editing protein TNFAIP3 and the NAD-dependent deacetylase sirtuin 1 (SIRT1). Interestingly, CM1 attenuated the Toll-like receptor 4 (TLR4)-induced production of inflammatory cytokines by inhibiting the extracellular-signal-regulated kinase (ERK)/MAPK and nuclear factor kappa B (NF-κB) signalling pathways. In addition, CM1 induced the expression of TNFAIP3 and SIRT1 on TLR4-stimulated primary macrophages; however, the anti-inflammatory effect of CM1 was abolished by the siRNA-mediated silencing of TNFAPI3 or by the genetic or pharmacologic inhibition of SIRT1. Importantly, intravenous administration of CM1 resulted in decreased susceptibility to endotoxin-induced sepsis, thereby attenuating the production of pro-inflammatory cytokines and neutrophil infiltration into the lung compared to control mice. Collectively, these findings demonstrate that CM1 has therapeutic potential for diverse inflammatory diseases, including sepsis.

The functional TNFAIP3 rs2230926T/G (Phe127Cys) variant confers risk to systemic lupus erythematosus in a Latin American population.

TNFAIP3 is a classical systemic lupus erythematosus (SLE)-associated risk locus identified by genome-wide association studies (GWASs) and replicated by candidate gene association studies primarily in Caucasians and Asians. However, in Latin American populations, its role on SLE susceptibility is not known. We conducted a case-control study to evaluate whether the TNFAIP3 rs2230926T/G (Phe127Cys) variant is associated with risk of developing SLE in a cohort of Mexican patients. The TNFAIP3 rs2230926T/G variant was analyzed in 561 patients with SLE and 499 control subjects, using TaqMan probes. We found that the G allele was associated with susceptibility to SLE under the allelic (OR 2.09, p = 0.005) and genotypic (OR 2.14, p = 0.004) models. In conclusion, our results show that TNFAIP3 rs2230926T/G is a risk factor for the development of SLE in the Mexican population.

[A20 haploinsufficiency: what do clinicians need to know?] / L'haploinsuffisance de A20 : que doit connaître le clinicien?

A20 Haploinsufficiency (HA20) is a monogenic autoinflammatory disease associated with an autosomal dominant mutation in the TNFAIP3 gene. It induces a defect in the inactivation of the pro-inflammatory NF-κB pathway. Less than 200 cases have been described worldwide. The clinical picture of the disease is essentially based on the association of recurrent fever and/or biologic inflammatory syndrome, aphtosis, often bipolar, and cutaneous folliculitis. However, the clinical spectrum of HA20 is very broad, including gastrointestinal (mainly colonic ulceration), articular, cutaneous, pericardial and lymph node involvement, as well as frequent association with organ-specific or non-specific autoimmune manifestations and/or autoantibodies, including antinuclear antibodies and anti-dsDNA. As a result, the diagnosis of a number of systemic or organic disorders, most notably Behçet's disease, Crohn's disease, and sometimes even systemic lupus, has been corrected to HA20 by molecular research for a heterozygous mutation with functional deficiency of TNFAIP3. Although the first signs of the disease often appear in the first years of life, the diagnosis is often made in adulthood and requires the involvement of both paediatric and adult physicians. Treatment for HA20 is not codified and relies on conventional or biological immunomodulators and immunosuppressants adapted to the patient's symptomatology. This review highlights the enormous diagnostic challenges in this autoinflammatory disease.

A20 Haploinsufficiency: A Systematic Review of 177 Cases.

A20 haploinsufficiency is an autoinflammatory disease caused by defective inactivation of the NF-κB pathway. We conducted a systematic literature review of articles reporting patients with TNFAIP3 sequence variants from 2016 to August 2023 following PRISMA (Preferred Reporting Items for Systematic Reviews and Meta-Analyses) guidelines. Data from 177 patients from 65 articles were retrieved (108 women). The principal features were mucosal ulcers (n = 129); fever (n = 93) followed by gastrointestinal (n = 81); skin features (n = 76); autoimmunity (n = 61), including thyroiditis (n = 25) and lupus (n = 16); and joint involvements (n = 54). Five patients had died at the time of publication. In 54 of 63 patients, CRP was significantly elevated during flares, with a median of 51 mg/l. The most commonly used treatment included corticosteroids and nonsteroidal anti-inflammatory drugs (n = 32), TNF blockers (n = 29), colchicine (n = 28), and methotrexate (n = 14). TNFAIP3 variants impacted the ovarian tumor domain in 92 cases and a Zinc finger domain in 68 cases. Geographic origin, reported sex, and variant type significantly impacted phenotype. A better understanding of the wide A20 haploinsufficiency phenotype could facilitate the diagnosis process. Much remains to be elucidated about pathogenesis and treatment to improve outcome in patients with A20 haploinsufficiency.

Advances in the SNPs of TNFAIP3 gene and autoimmune diseases / 国际儿科学杂志

The gene TNFAIP3 acts as a negative regulator of the NF-κB signaling pathway. TNFAIP3 encodes the A20 protein，which exerts a potent anti-inflammatory effect and plays a pivotal role in the regulation of inflammation and immunity. In recent years，TNFAIP3 has garnered significant attention as a susceptibility gene for numerous autoimmune diseases，including but not limited to systemic lupus erythematosus，rheumatoid arthritis，psoriasis. Additionally，high-penetrance heterozygous mutations in TNFAIP3 cause a haploinsufficiency of A20（HA20）. HA20 is a monogenic autoinflammatory disease. But some individuals of HA20 exhibit clinical features of autoimmune diseases，including varying degrees of autoantibody positivity，lupus-like phenotypes，and autoimmune thyroid disease.This article focuses on the single nucleotide polymorphism of TNFAIP3 and related autoimmune diseases，to underscore the crucial role of TNFAIP3 in the pathogenesis of autoimmune diseases，and to provide new research directions and potential drug targets for these conditions.

Protective effect of TNFAIP3 on testosterone production in Leydig cells under an aging inflammatory microenvironment.

BACKGROUND: The aging inflammatory microenvironment surrounding Leydig cells is linked to reduced testosterone levels in males. Tumor necrosis factor alpha-induced protein 3 (TNFAIP3) acts as a critical anti-inflammatory factor in various aging-related diseases. This study aims to investigate the protective effect of TNFAIP3 on testosterone production in Leydig cells under an aging inflammatory microenvironment. METHODS: Bioinformatics analysis examined TNFAIP3 expression differences in aging rat testes and validated the findings in aging mouse testes. In vitro models of inflammation were established using two Leydig cell lines, with tumor necrosis factor alpha (TNF-α) as the inflammatory factor. Lentiviral transduction was utilized to manipulate TNFAIP3 expression in these cell lines. Transcriptomic sequencing identified differentially expressed genes in TNFAIP3-overexpressing cells. RESULTS: Bioinformatics analysis and validation experiments revealed increased inflammatory signaling and elevated TNFAIP3 expression in aging rat and mouse testes. TNFAIP3 knockdown worsened testosterone synthesis inhibition and apoptosis in cells, while TNFAIP3 overexpression reversed these effects. Transcriptome analysis identified alterations in the P38MAPK pathway following TNFAIP3 overexpression. TNFAIP3 knockdown enhanced TNF-induced P38MAPK signaling, whereas its overexpression attenuated this effect. TNFAIP3 was found to regulate testosterone synthesis by upregulating CEBPB expression. CONCLUSIONS: TNFAIP3 exhibits inhibitory effects on apoptosis and promotes testosterone production in Leydig cells. The protective influence of TNFAIP3 on Leydig cells within an inflammatory microenvironment is likely mediated through by inhibiting the P38MAPK pathway and upregulating CEBPB expression.

Genomic associations with antibody response to an oral cholera vaccine.

Oral cholera vaccine is one of the key interventions used in our fight to end the longest pandemic of our time, cholera. The immune response conferred by the currently available cholera vaccines, as measured by serum antibody levels, is variable amongst its recipients. We undertook a genome wide association study (GWAS) on antibody response to the cholera vaccine; globally, the first GWAS on cholera vaccine response. We identified three clusters of bi-allelic SNPs, in high within-cluster linkage disequilibrium that were moderately (p < 5 × 10-6) associated with antibody response to the cholera vaccine and mapped to chromosomal regions 4p14, 4p16.1 and 6q23.3. Intronic SNPs of TBC1D1 comprised the cluster on 4p14, intronic SNPs of TBC1D14 comprised that on 4p16.1 and SNPs upstream of TNFAIP3 formed the cluster on 6q23.3. SNPs within and around these clusters have been implicated in immune cell function and immunological aspects of autoimmune or infectious diseases (e.g., diseases caused by Helicobacter pylori and malarial parasite). 6q23.3 is a prominent region harbouring many loci associated with immune related diseases, including multiple sclerosis, rheumatoid arthritis and systemic lupus erythematosus, as well as IL2 and INFα response to a smallpox vaccine. The gene clusters identified in this study play roles in vesicle-mediated pathway, autophagy and NF-κB signaling. No significant effect of O blood group on antibody response to the cholera vaccine was observed in this study.

Haploinsufficiency of A20 caused by a novel pathogenic missense variant of TNFAIP3 and successfully treated with anti-TNF and immunosuppressive therapies.

Loss-of-function of protein A20, encoded by TNFAIP3, leads to an early-onset haploinsufficiency of A20 (HA20). This study reports one Chinese child with HA20 and explores the genetic etiology of TNFAIP3 variant. The patient exhibited transient recurrent episodes of fever, intermittent signs of arthritis, gastrointestinal symptoms and multiple colonic ulcers. Laboratory tests revealed elevated inflammatory indicators and mild to moderate anemia. Genetic analysis identified a heterozygous de novo variant in his TNFAIP3 gene (c.740C＞T, p. P247L), which had never been reported before. The novel missense variation was validated to be pathogenic through causing insufficient expression of A20, over-activation of NF-κB signaling pathway and elevated levels of proinflammatory cytokines in response to stimulation by lipopolysaccharide. A combination of oral corticosteroids, TNF-α inhibitors and thalidomide freed him from symptoms and abnormal inflammatory indicators. Furthermore, continual improvement of the patient's condition was observed during a follow-up period of five months. We demonstrate a case with a de novo missense variant resulting in a loss-of-function of TNFAIP3, which expands the clinical spectrum of HA20. Cytokine antagonists and immunosuppressants may be effective drugs.