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Kawasaki disease (KD) is the leading cause of acquired heart disease among children. Murine and human data suggest that the NLRP3-IL-1β pathway is the main driver of KD pathophysiology. NLRP3 can be activated during defective autophagy/mitophagy. We used the Lactobacillus casei cell wall extract (LCWE) murine model of KD vasculitis to examine the role of autophagy/mitophagy on cardiovascular lesion development. LCWE-injected mice had impaired autophagy/mitophagy and increased levels of ROS in cardiovascular lesions, together with increased systemic 8-OHdG release. Enhanced autophagic flux significantly reduced cardiovascular lesions in LCWE-injected mice, whereas autophagy blockade increased inflammation. Vascular smooth muscle cell-specific deletion of Atg16l1 and global Parkin-/- significantly increased disease formation, supporting the importance of autophagy/mitophagy in this model. Ogg1-/- mice had significantly increased lesions with increased NLRP3 activity, whereas treatment with MitoQ reduced vascular tissue inflammation, ROS production, and systemic 8-OHdG release. Treatment with MN58b or Metformin (increasing AMPK and reducing ROS) resulted in decreased cardiovascular lesions. Our results demonstrate that impaired autophagy/mitophagy and ROS-dependent damage exacerbate the development of murine KD vasculitis. This pathway can be efficiently targeted to reduce disease severity. These findings enhance our understanding of KD pathogenesis and identify potentially novel therapeutic avenues for KD treatment.

We recently discovered a superantigen-like motif sequentially and structurally similar to a staphylococcal enterotoxin B (SEB) segment, near the S1/S2 cleavage site of the SARS-CoV-2 spike protein, which might explain the multisystem inflammatory syndrome (MIS-C) observed in children and the cytokine storm in severe COVID-19 patients. We show here that an anti-SEB monoclonal antibody (mAb), 6D3, can bind this viral motif at its polybasic (PRRA) insert to inhibit infection in live virus assays. The overlap between the superantigenic site of the spike and its proteolytic cleavage site suggests that the mAb prevents viral entry by interfering with the proteolytic activity of cell proteases (furin and TMPRSS2). The high affinity of 6D3 for this site originates from a polyacidic segment at its heavy chain CDR2. The study points to the potential utility of 6D3 for possibly treating COVID-19, MIS-C, or common colds caused by human coronaviruses that also possess a furin-like cleavage site.

KD is an acute febrile illness and systemic vasculitis of unknown etiology among young children, which can cause coronary artery abnormalities and aneurysms (CAA) and is the leading cause of acquired heart disease among children in the US. Lactobacillus casei cell wall extract (LCWE) induces in mice a vasculitis following intraperitoneal injection defined by the activation of macrophages, dendritic cells and CD8+ cytotoxic T cells leading to aortitis, coronary arteritis, aneurysms and myocarditis that strongly mimic the immunopathology and the cardiac lesions observed in children with Kawasaki disease (KD). To address a potential pathogenic role of LCWE-specific T cells in human vascular inflammation, we studied the activation of circulating CD4+ and CD8+ T cells ex vivo in response to LCWE in 3 cohorts: (1) KD children 2-3 weeks after fever onset, (2) age-similar healthy children controls, (3) healthy adult controls. In all subjects studied, pro-inflammatory CD4+ and CD8+T cells responded to LCWE with no significant differences. Peripherally-induced regulatory T cells (iTreg) also responded to LCWE and potentially reverted to Th17, as suggested by the detection of IL-17 in culture supernatants. Central memory T cells were also detectable and were more abundant in adults. The potential homing to the vessels of LCWE-specific T cells was suggested by the expression of CCR6 and CD31. In conclusion, a non-pathogenic, LCWE-specific T cell repertoire could lead to KD depending upon priming conditions, genetic factors and immune activation by other antigens.

Receptor interacting protein 2 (RIP2) plays a role in sensing intracellular pathogens, but its function in T cells is unclear. We show that RIP2 deficiency in CD4+ T cells resulted in chronic and severe interleukin-17A-mediated inflammation during Chlamydia pneumoniae lung infection, increased T helper 17 (Th17) cell formation in lungs of infected mice, accelerated atherosclerosis, and more severe experimental autoimmune encephalomyelitis. While RIP2 deficiency resulted in reduced conventional Th17 cell differentiation, it led to significantly enhanced differentiation of pathogenic (p)Th17 cells, which was dependent on RORα transcription factor and interleukin-1 but independent of nucleotide oligomerization domain (NOD) 1 and 2. Overexpression of RIP2 resulted in suppression of pTh17 cell differentiation, an effect mediated by its CARD domain, and phenocopied by a cell-permeable RIP2 CARD peptide. Our data suggest that RIP2 has a T cell-intrinsic role in determining the balance between homeostatic and pathogenic Th17 cell responses.

ObjectiveKawasaki disease (KD) is the leading cause of acquired heart disease among children in developed countries. Coronary lesions in KD in humans are characterized by an increased presence of infiltrating CD3+ T cells; however, the specific contributions of the different T cell subpopulations in coronary arteritis development remain unknown. Therefore, we sought to investigate the function of CD4+ and CD8+ T cells, Treg cells, and natural killer (NK) T cells in the pathogenesis of KD.MethodsWe addressed the function of T cell subsets in KD development by using a well-established murine model of Lactobacillus casei cell wall extract (LCWE)-induced KD vasculitis. We determined which T cell subsets were required for development of KD vasculitis by using several knockout murine strains and depleting monoclonal antibodies.ResultsLCWE-injected mice developed coronary lesions characterized by the presence of inflammatory cell infiltrates. Frequently, this chronic inflammation resulted in complete occlusion of the coronary arteries due to luminal myofibroblast proliferation (LMP) as well as the development of coronary arteritis and aortitis. We found that CD8+ T cells, but not CD4+ T cells, NK T cells, or Treg cells, were required for development of KD vasculitis.ConclusionThe LCWE-induced murine model of KD vasculitis mimics many histologic features of the disease in humans, such as the presence of CD8+ T cells and LMP in coronary artery lesions as well as epicardial coronary arteritis. Moreover, CD8+ T cells functionally contribute to the development of KD vasculitis in this murine model. Therapeutic strategies targeting infiltrating CD8+ T cells might be useful in the management of KD in humans.

Multisystem inflammatory syndrome in children (MIS-C) is a rare pediatric inflammatory disorder characterized by immune cell hyperactivation, cytokine storm, and the production of autoantibodies. The mechanisms underlying such immune dysregulation still need to be unraveled. In this issue of theJCI, Benamar et al. demonstrated the critical role of the Notch receptor 1/CD22 (Notch1/CD22) axis in Tregs, which, when activated, impairs Treg functions and promotes inflammation. They showed that the Notch1/CD22 axis contributed to dysregulated immune responses in MIS-C. These findings may have implications for MIS-C and many other inflammatory diseases., Multisystem inflammatory syndrome in children In adults, COVID-19 can result in severe and potentially lethal respiratory complications associated with dysregulated and robust inflammatory responses. Children with COVID-19 are usually asymptomatic [...]

Multisystem inflammatory syndrome in children (MIS-C) manifests as a severe and uncontrolled inflammatory response with multiorgan involvement, occurring weeks after SARS-CoV-2 infection. Here, we utilized proteomics, RNA sequencing, autoantibody arrays, and B cell receptor (BCR) repertoire analysis to characterize MIS-C immunopathogenesis and identify factors contributing to severe manifestations and intensive care unit admission. Inflammation markers, humoral immune responses, neutrophil activation, and complement and coagulation pathways were highly enriched in MIS-C patient serum, with a more hyperinflammatory profile in severe than in mild MIS-C cases. We identified a strong autoimmune signature in MIS-C, with autoantibodies targeted to both ubiquitously expressed and tissue-specific antigens, suggesting autoantigen release and excessive antigenic drive may result from systemic tissue damage. We further identified a cluster of patients with enhanced neutrophil responses as well as high anti-Spike IgG and autoantibody titers. BCR sequencing of these patients identified a strong imprint of antigenic drive with substantial BCR sequence connectivity and usage of autoimmunity-associated immunoglobulin heavy chain variable region (IGHV) genes. This cluster was linked to a TRBV112 expanded T cell receptor (TCR) repertoire, consistent with previous studies indicating a superantigen-driven pathogenic process. Overall, we identify a combination of pathogenic pathways that culminate in MIS-C and may inform treatment., Introduction While cases of severe coronavirus disease 2019 (COVID-19) occur relatively less frequently in children than adults (1, 2), a small proportion of SARS-CoV-2-infected children develop a novel pediatric febrile [...]

BACKGROUND. Weeks after SARS-CoV-2 infection or exposure, some children develop a severe, life-threatening illness called multisystem inflammatory syndrome in children (MIS-C). Gastrointestinal (GI) symptoms are common in patients with MIS-C, and a severe hyperinflammatory response ensues with potential for cardiac complications. The cause of MIS-C has not been identified to date. METHODS. Here, we analyzed biospecimens from 100 children: 19 with MIS-C, 26 with acute COVID-19, and 55 controls. Stools were assessed for SARS-CoV-2 by reverse transcription PCR (RT-PCR), and plasma was examined for markers of breakdown of mucosal barrier integrity, including zonulin. Ultrasensitive antigen detection was used to probe for SARS-CoV-2 antigenemia in plasma, and immune responses were characterized. As a proof of concept, we treated a patient with MIS-C with larazotide, a zonulin antagonist, and monitored the effect on antigenemia and the patient's clinical response. RESULTS. We showed that in children with MIS-C, a prolonged presence of SARS-CoV-2 in the GI tract led to the release of zonulin, a biomarker of intestinal permeability, with subsequent trafficking of SARS-CoV-2 antigens into the bloodstream, leading to hyperinflammation. The patient with MIS-C treated with larazotide had a coinciding decrease in plasma SARSCoV-2 spike antigen levels and inflammatory markers and a resultant clinical improvement above that achieved with currently available treatments. CONCLUSION. These mechanistic data on MIS-C pathogenesis provide insight into targets for diagnosing, treating, and preventing MIS-C, which are urgently needed for this increasingly common severe COVID-19-related disease in children., Introduction Most children who are acutely infected with SARS-CoV-2 develop mild upper respiratory symptoms or experience asymptomatic infection. Several days to weeks after resolution of the initial infection, some of [...]

Multisystem inflammatory syndrome in children (MIS-C), a hyperinflammatory syndrome associated with SARS-CoV-2 infection, shares clinical features with toxic shock syndrome, which is triggered by bacterial superantigens. Superantigen specificity for different V[beta] chains results in V[beta] skewing, whereby T cells with specific V[beta] chains and diverse antigen specificity are overrepresented in the T cell receptor (TCR) repertoire. Here, we characterized the TCR repertoire of MIS-C patients and found a profound expansion of TCR[beta] variable gene 11-2 (TRBV11-2), with up to 24% of clonal T cell space occupied by TRBV112 T cells, which correlated with MIS-C severity and serum cytokine levels. Analysis of TRBJ gene usage and complementarity-determining region 3 (CDR3) length distribution of MIS-C expanded TRBV11-2 clones revealed extensive junctional diversity. Patients with TRBV11-2 expansion shared HLA class I alleles A02, B35, and C04, indicating what we believe is a novel mechanism for CDR3-independent T cell expansion. In silico modeling indicated that polyacidic residues in the V[beta] chain encoded by TRBV11-2 (V[beta]21.3) strongly interact with the superantigen- like motif of SARS-CoV-2 spike glycoprotein, suggesting that unprocessed SARS-CoV-2 spike may directly mediate TRBV11-2 expansion. Overall, our data indicate that a CDR3-independent interaction between SARS-CoV-2 spike and TCR leads to T cell expansion and possibly activation, which may account for the clinical presentation of MIS-C., Introduction Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of coronavirus disease 2019 (COVID-19), which started as an epidemic in China and culminated in a global pandemic. [...]

BACKGROUND. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused more than 1 million deaths worldwide; thus, there is an urgent need to develop preventive and therapeutic strategies. The antituberculosis vaccine bacillus Calmette-Guerin (BCG) demonstrates nonspecific, protective innate immune-boosting effects. Here, we determined whether a history of BCG vaccination was associated with decreased SARS-CoV-2 infection and seroconversion in a longitudinal, retrospective observational study of a diverse cohort of health care workers (HCWs). METHODS. We assessed SARS-CoV-2 seroprevalence and collected medical questionnaires, which included information on BCG vaccination status and preexisting demographic and clinical characteristics, from an observational cohort of HCWs in a multisite Los Angeles health care organization. We used multivariate analysis to determine whether a history of BCG vaccination was associated with decreased rates of SARS-CoV-2 infection and seroconversion. RESULTS. Of the 6201 HCWs, 29.6% reported a history of BCG vaccination, whereas 68.9% had not received BCG vaccination. Seroprevalence of anti-SARS-CoV-2 IgG as well as the incidence of self-reported clinical symptoms associated with coronavirus disease 2019 (COVID-19) were markedly decreased among HCWs with a history of BCG vaccination compared with those without BCG vaccination. After adjusting for age and sex, we found that a history of BCG vaccination, but not meningococcal, pneumococcal, or influenza vaccination, was associated with decreased SARS-CoV-2 IgG seroconversion. CONCLUSIONS. A history of BCG vaccination was associated with a decrease in the seroprevalence of anti-SARS-CoV-2 IgG and a lower number of participants who self-reported experiencing COVID-19-related clinical symptoms in this cohort of HCWs. Therefore, large randomized, prospective clinical trials of BCG vaccination are urgently needed to confirm whether BCG vaccination can confer a protective effect against SARS-CoV-2 infection., Introduction As of early October 2020, more than 37.5 million people worldwide had been infected by the novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which has resulted in more [...]

Tregs expressing the transcription factor Foxp3 suppress self-reactive T cells, prevent autoimmunity, and help contain immune responses to foreign antigens, thereby limiting the potential for inadvertent tissue damage. Mutations in the FOXP3 gene result in Treg deficiency in mice and humans, which leads to the development of a multisystem autoimmune inflammatory disease. The contribution of dysregulated innate immune responses to the pathogenesis of Foxp3 deficiency disease is unknown. In this study, we examined the role of microbial signals in the pathogenesis of Foxp3 deficiency disease by studying Foxp3 mutant mice that had concurrent deficiencies in TLR signaling pathways. Global deficiency of the common TLR adaptor MyD88 offered partial protection from Foxp3 deficiency disease. Specifically, it protected from disease at the environmental inter-faces of the skin, lungs, and gut. In contrast, systemic disease, in the form of unrestrained lymphoproliferation, continued unabated. The effect of MyD88 deficiency at environmental interfaces involved the disruption of chemokine gradients that recruit effector T cells and DCs, resulting in their entrapment in secondary lymphoid tissues. These results suggests that Tregs have a key role in maintaining tolerance at host-microbial interfaces by restraining tonic MyD88-dependent proinflammatory signals. Moreover, microbial factors may play a substantial role in the pathogenesis of human autoimmune disease resulting from Treg deficiency., Introduction Foxp[3.sup.+] Tregs are central to the maintenance of peripheral tolerance (1). Tregs exert dominant suppression of self-reactive T cells and help contain immune responses to foreign antigens, thereby limiting [...]

Multisystem inflammatory syndrome in children (MIS-C) is a febrile pediatric inflammatory disease that may develop weeks after initial SARS-CoV-2 infection or exposure. MIS-C involves systemic hyperinflammation and multiorgan involvement, including severe cardiovascular, gastrointestinal (GI) and neurological symptoms. Some clinical attributes of MIS-C--such as persistent fever, rashes, conjunctivitis and oral mucosa changes (red fissured lips and strawberry tongue)--overlap with features of Kawasaki disease (KD). In addition, MIS-C shares striking clinical similarities with toxic shock syndrome (TSS), which is triggered by bacterial superantigens (SAgs). The remarkable similarities between MIS-C and TSS prompted a search for SAg-like structures in the SARS-CoV-2 virus and the discovery of a unique SAg-like motif highly similar to a Staphylococcal enterotoxin B (SEB) fragment in the SARS-CoV-2 spike 1 (S1) glycoprotein. Computational studies suggest that the SAg-like motif has a high affinity for binding T-cell receptors (TCRs) and MHC Class II proteins. Immunosequencing of peripheral blood samples from MIS-C patients revealed a profound expansion of TCR b variable gene 11-2 (TRBV11-2), which correlates withMIS-C severity and serum cytokine levels, consistent with a SAg-triggered immune response. Computational sequence analysis of SARS-CoV-2 spike further identified conserved neurotoxin-like motifs which may alter neuronal cell function and contribute to neurological symptoms in COVID-19 and MIS-C patients. Additionally, autoantibodies are detected during MIS-C, which may indicate development of post-SARS-CoV-2 autoreactive and autoimmune responses. Finally, prolonged persistence of SARS-CoV-2 RNA in the gut, increased gut permeability and elevated levels of circulating S1 have been observed in children with MIS-C. Accordingly, we hypothesize that continuous and prolonged exposure to the viral SAg-like and neurotoxin-like motifs in SARS-CoV-2 spike may promote autoimmunity leading to the development of postacute COVID-19 syndromes, including MIS-C and long COVID, as well as the neurological complications resulting from SARS-CoV-2 infection. [ABSTRACT FROM AUTHOR]

Immunoglobulin E (IgE) antibodies are known for triggering immediate hypersensitivity reactions such as food anaphylaxis. In this study, we tested whether they might additionally function to amplify nascent antibody and T helper 2 (Th2) cell-mediated responses to ingested proteins and whether blocking IgE would modify sensitization. By using mice harboring a disinhibited form of the IL-4 receptor, we developed an adjuvant-free model of peanut allergy. Mast cells and IgE were required for induction of antibody and Th2-cell-mediated responses to peanut ingestion and they impaired regulatory T (Treg) cell induction. Mast-cell-targeted genetic deletion of the FcεRI signaling kinase Syk or Syk blockade also prevented peanut sensitization. In mice with established allergy, Syk blockade facilitated desensitization and induction of Treg cells, which suppressed allergy when transferred to naive recipients. Our study suggests a key role for IgE in driving Th2 cell and IgE responses while suppressing Treg cells in food allergy. [ABSTRACT FROM AUTHOR]

Alterations in the intestinal microbiota contribute to the pathogenesis of various cardiovascular disorders, but how they affect the development of Kawasaki disease (KD), an acute pediatric vasculitis, remains unclear. We report that depleting the gut microbiota reduces the development of cardiovascular inflammation in a murine model mimicking KD vasculitis. The development of cardiovascular lesions was associated with alterations in the intestinal microbiota composition and, notably, a decreased abundance of Akkermansia muciniphila and Faecalibacterium prausnitzii. Oral supplementation with either of these live or pasteurized individual bacteria, or with short-chain fatty acids (SCFAs) produced by them, attenuated cardiovascular inflammation. Treatment with Amuc_1100, the TLR-2 signaling outer membrane protein from A. muciniphila , also decreased the severity of vascular inflammation. This study reveals an underappreciated gut microbiota-cardiovascular inflammation axis in KD vasculitis pathogenesis and identifies specific intestinal commensals that regulate vasculitis in mice by producing metabolites or via extracellular proteins acting on gut barrier function., In Brief: It remains unclear whether changes in the intestinal microbiota composition are involved in the development of cardiovascular lesions associated with Kawasaki disease (KD), an immune-mediated vasculitis. Jena et al. observe alterations in the intestinal microbiota composition of mice developing vasculitis, characterized by reduced A. muciniphila and F. prausnitzii . Oral supplementation with either of these bacteria, live or pasteurized, or with bacteria-produced short-chain fatty acids (SCFAs) or Amuc_1100, the TLR-2 signaling outer membrane protein of A. muciniphila , was sufficient to alleviate the development of cardiovascular lesions in mice by promoting intestinal barrier function., Highlights: Absence or depletion of the microbiota decreases the severity of vasculitis in a murine model mimicking KD vasculitis. Supplementation of B. wadsworthia and B. fragilis promotes murine KD vasculitis. Decreased abundances of F. prausnitzii and A. muciniphila are associated with the development of cardiovascular lesions in mice. Supplementation with either live or pasteurized A. muciniphila and F. prausnitzii, or the TLR-2 signaling Amuc_1100, reduces the severity of vasculitis by promoting gut barrier function.