Mast cells mediate neutrophil recruitment and vascular leakage through the NLRP3 inflammasome in histamine-independent urticaria

Urticarial rash observed in cryopyrin-associated periodic syndrome (CAPS) caused by nucleotide-binding oligomerization domain–leucine-rich repeats containing pyrin domain 3 (NLRP3) mutations is effectively suppressed by anti–interleukin (IL)-1 treatment, suggesting a pathophysiological role of IL-1β in the skin. However, the cellular mechanisms regulating IL-1β production in the skin of CAPS patients remain unclear. We identified mast cells (MCs) as the main cell population responsible for IL-1β production in the skin of CAPS patients. Unlike normal MCs that required stimulation with proinflammatory stimuli for IL-1β production, resident MCs from CAPS patients constitutively produced IL-1β. Primary MCs expressed inflammasome components and secreted IL-1β via NLRP3 and apoptosis-associated speck-like protein containing a caspase recruitment domain when stimulated with microbial stimuli known to activate caspase-1. Furthermore, MCs expressing disease-associated but not wild-type NLRP3 secreted IL-1β and induced neutrophil migration and vascular leakage, the histological hallmarks of urticarial rash, when transplanted into mouse skin. Our findings implicate MCs as IL-1β producers in the skin and mediators of histamine-independent urticaria through the NLRP3 inflammasome

Trapping of CDC42 C-terminal variants in the Golgi drives pyrin inflammasome hyperactivation

CDC42-C末端異常症に於ける炎症病態を解明 --ゴルジ体への異常蓄積がパイリンインフラマソーム形成を過剰促進--. 京都大学プレスリリース. 2022-05-02.Mutations in the C-terminal region of the CDC42 gene cause severe neonatal-onset autoinflammation. Effectiveness of IL-1β–blocking therapy indicates that the pathology involves abnormal inflammasome activation; however, the mechanism underlying autoinflammation remains to be elucidated. Using induced-pluripotent stem cells established from patients carrying CDC42[R186C], we found that patient-derived cells secreted larger amounts of IL-1β in response to pyrin-activating stimuli. Aberrant palmitoylation and localization of CDC42[R186C] protein to the Golgi apparatus promoted pyrin inflammasome assembly downstream of pyrin dephosphorylation. Aberrant subcellular localization was the common pathological feature shared by CDC42 C-terminal variants with inflammatory phenotypes, including CDC42[*192C*24] that also localizes to the Golgi apparatus. Furthermore, the level of pyrin inflammasome overactivation paralleled that of mutant protein accumulation in the Golgi apparatus, but not that of the mutant GTPase activity. These results reveal an unexpected association between CDC42 subcellular localization and pyrin inflammasome activation that could pave the way for elucidating the mechanism of pyrin inflammasome formation

STING signalling is terminated through ESCRT-dependent microautophagy of vesicles originating from recycling endosomes

STING炎症シグナルの終結分子機構 --新規細胞内分解システムの発見--. 京都大学プレスリリース. 2023-03-14.Stimulator of interferon genes (STING) is essential for the type I interferon response against a variety of DNA pathogens. Upon emergence of cytosolic DNA, STING translocates from the endoplasmic reticulum to the Golgi where STING activates the downstream kinase TBK1, then to lysosome through recycling endosomes (REs) for its degradation. Although the molecular machinery of STING activation is extensively studied and defined, the one underlying STING degradation and inactivation has not yet been fully elucidated. Here we show that STING is degraded by the endosomal sorting complexes required for transport (ESCRT)-driven microautophagy. Airyscan super-resolution microscopy and correlative light/electron microscopy suggest that STING-positive vesicles of an RE origin are directly encapsulated into Lamp1-positive compartments. Screening of mammalian Vps genes, the yeast homologues of which regulate Golgi-to-vacuole transport, shows that ESCRT proteins are essential for the STING encapsulation into Lamp1-positive compartments. Knockdown of Tsg101 and Vps4, components of ESCRT, results in the accumulation of STING vesicles in the cytosol, leading to the sustained type I interferon response. Knockdown of Tsg101 in human primary T cells leads to an increase the expression of interferon-stimulated genes. STING undergoes K63-linked ubiquitination at lysine 288 during its transit through the Golgi/REs, and this ubiquitination is required for STING degradation. Our results reveal a molecular mechanism that prevents hyperactivation of innate immune signalling, which operates at REs

ISSAID/EMQN Best Practice Guidelines for the Genetic Diagnosis of Monogenic Autoinflammatory Diseases in the Next-Generation Sequencing Era

Abstract Background Monogenic autoinflammatory diseases are caused by pathogenic variants in genes that regulate innate immune responses, and are characterized by sterile systemic inflammatory episodes. Since symptoms can overlap within this rapidly expanding disease category, accurate genetic diagnosis is of the utmost importance to initiate early inflammation-targeted treatment and prevent clinically significant or life-threatening complications. Initial recommendations for the genetic diagnosis of autoinflammatory diseases were limited to a gene-by-gene diagnosis strategy based on the Sanger method, and restricted to the 4 prototypic recurrent fevers (MEFV, MVK, TNFRSF1A, and NLRP3 genes). The development of best practices guidelines integrating critical recent discoveries has become essential. Methods The preparatory steps included 2 online surveys and pathogenicity annotation of newly recommended genes. The current guidelines were drafted by European Molecular Genetics Quality Network members, then discussed by a panel of experts of the International Society for Systemic Autoinflammatory Diseases during a consensus meeting. Results In these guidelines, we combine the diagnostic strength of next-generation sequencing and recommendations to 4 more recently identified genes (ADA2, NOD2, PSTPIP1, and TNFAIP3), nonclassical pathogenic genetic alterations, and atypical phenotypes. We present a referral-based decision tree for test scope and method (Sanger versus next-generation sequencing) and recommend on complementary explorations for mosaicism, copy-number variants, and gene dose. A genotype table based on the 5-category variant pathogenicity classification provides the clinical significance of prototypic genotypes per gene and disease. Conclusions These guidelines will orient and assist geneticists and health practitioners in providing up-to-date and appropriate diagnosis to their patients

Controlo químico de infestantes

Uma planta é considerada infestante quando nasce espontaneamente num local e momento indesejados, podendo interferir negativamente com a cultura instalada. As infestantes competem com as culturas para o espaço, a luz, água e nutrientes, podendo atrasar e prejudicar as operações de colheita, depreciar o produto final e assegurarem a reinfestação nas culturas seguintes. Dado o modo de propagação diferenciado das diversas espécies de infestantes, com as anuais a propagarem-se por semente e as perenes ou vivazes a assegurarem a sua propagação através de órgãos vegetativos (rizomas, bolbos, tubérculos, etc.), assim, também o seu controlo quer químico, quer mecânico terá que ser diferenciado, ou seja, para controlar infestantes anuais será suficiente destruir a sua parte aérea, enquanto para controlar infestantes perenes teremos que destruir os seus órgãos reprodutivos. O controlo de infestantes poderá ser químico, através da utilização de herbicidas, ou mecânico pela utilização de alfaias agrícolas, tais como a charrua de aivecas, a charrua de discos, a grade de discos, o escarificador e a fresa. Quando a técnica utilizada na instalação das culturas é a sementeira directa, o controlo das infestantes terá que ser obrigatoriamente químico, enquanto se o recurso à mobilização do solo for a técnica mais utilizada (sistema de mobilização tradicional ou sistema de mobilização reduzida), o controlo das infestantes tanto poderá ser químico como mecânico. Neste trabalho iremos abordar apenas, o controlo químico de infestantes

Somatic NLRP3 mosaicism in Muckle-Wells syndrome. A genetic mechanism shared by different phenotypes of cryopyrin-associated periodic syndromes

Familial cold autoinflammatory syndrome, Muckle-Wells syndrome (MWS), and chronic, infantile, neurological, cutaneous and articular (CINCA) syndrome are dominantly inherited autoinflammatory diseases associated to gain-of-function NLRP3 mutations and included in the cryopyrin-associated periodic syndromes (CAPS). A variable degree of somatic NLRP3 mosaicism has been detected in ≈35% of patients with CINCA. However, no data are currently available regarding the relevance of this mechanism in other CAPS phenotypes. OBJECTIVE: To evaluate somatic NLRP3 mosaicism as the disease-causing mechanism in patients with clinical CAPS phenotypes other than CINCA and NLRP3 mutation-negative. METHODS: NLRP3 analyses were performed by Sanger sequencing and by massively parallel sequencing. Apoptosis-associated Speck-like protein containing a CARD (ASC)-dependent nuclear factor kappa-light chain-enhancer of activated B cells (NF-κB) activation and transfection-induced THP-1 cell death assays determined the functional consequences of the detected variants. RESULTS: A variable degree (5.5-34.9%) of somatic NLRP3 mosaicism was detected in 12.5% of enrolled patients, all of them with a MWS phenotype. Six different missense variants, three novel (p.D303A, p.K355T and p.L411F), were identified. Bioinformatics and functional analyses confirmed that they were disease-causing, gain-of-function NLRP3 mutations. All patients treated with anti-interleukin1 drugs showed long-lasting positive responses. CONCLUSIONS: We herein show somatic NLRP3 mosaicism underlying MWS, probably representing a shared genetic mechanism in CAPS not restricted to CINCA syndrome. The data here described allowed definitive diagnoses of these patients, which had serious implications for gaining access to anti-interleukin 1 treatments under legal indication and for genetic counselling. The detection of somatic mosaicism is difficult when using conventional methods. Potential candidates should benefit from the use of modern genetic tool

Diagnosis and Treatment in Anhidrotic Ectodermal Dysplasia with Immunodeficiency

ABSTRACTAnhidrotic ectodermal dysplasia with immunodeficiency (EDA-ID) is characterized according to its various manifestations, which include ectodermal dysplasia, vascular anomalies, osteopetrosis, and diverse immunological abnormalities such as susceptibility to pathogens, impaired antibody responses to polysaccharides, hypogammaglobulinemia, hyper-IgM syndrome, impaired natural killer cell cytotoxicity, and autoimmune diseases. Two genes responsible for EDA-ID have been identified: nuclear factor-κB (NF-κB) essential modulator (NEMO) for X-linked EDA-ID (XL-EDA-ID) and IκBα for autosomal-dominant EDA-ID (AD-EDA-ID). Both genes are involved in NF-κB activation, such that mutations or related defects cause impaired NF-κB signaling. In particular, NEMO mutations are scattered across the entire NEMO gene in XL-EDA-ID patients, which explains the broad spectrum of clinical manifestations and the difficulties associated with making a diagnosis. In this review, we focus on the pathophysiology of EDA-ID and different diagnostic strategies, which will be beneficial for early diagnosis and appropriate treatment