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Abstract Acquired mutations in the UBA1 gene were recently identified in patients with severe adult-onset auto-inflammatory syndrome called VEXAS (vacuoles, E1 enzyme, X-linked, autoinflammatory, somatic). However, the precise physiological and clinical impact of these mutations remains poorly defined. Here we study a unique prospective cohort of VEXAS patients. We show that monocytes from VEXAS are quantitatively and qualitatively impaired and display features of exhaustion with aberrant expression of chemokine receptors. In peripheral blood from VEXAS patients, we identify an increase in circulating levels of many proinflammatory cytokines, including IL-1β and IL-18 which reflect inflammasome activation and markers of myeloid cells dysregulation. Gene expression analysis of whole blood confirms these findings and also reveals a significant enrichment of TNF-α and NFκB signaling pathways that can mediate cell death and inflammation. This study suggests that the control of the nflammasome activation and inflammatory cell death could be therapeutic targets in VEXAS syndrome.

Abstract Loss of NBEAL2 function leads to grey platelet syndrome (GPS), a bleeding disorder characterized by macro-thrombocytopenia and α-granule-deficient platelets. A proportion of patients with GPS develop autoimmunity through an unknown mechanism, which might be related to the proteins NBEAL2 interacts with, specifically in immune cells. Here we show a comprehensive interactome of NBEAL2 in primary T cells, based on mass spectrometry identification of altogether 74 protein association partners. These include LRBA, a member of the same BEACH domain family as NBEAL2, recessive mutations of which cause autoimmunity and lymphocytic infiltration through defective CTLA-4 trafficking. Investigating the potential association between NBEAL2 and CTLA-4 signalling suggested by the mass spectrometry results, we confirm by co-immunoprecipitation that CTLA-4 and NBEAL2 interact with each other. Interestingly, NBEAL2 deficiency leads to low CTLA-4 expression in patient-derived effector T cells, while their regulatory T cells appear unaffected. Knocking-down NBEAL2 in healthy primary T cells recapitulates the low CTLA-4 expression observed in the T cells of GPS patients. Our results thus show that NBEAL2 is involved in the regulation of CTLA-4 expression in conventional T cells and provide a rationale for considering CTLA-4-immunoglobulin therapy in patients with GPS and autoimmune disease.

BackgroundIn most cases, Zika virus (ZIKV) causes a self-limited acute illness in adults, characterized by mild clinical symptoms that resolve within a few days. Immune responses, both innate and adaptive, play a central role in controlling and eliminating virus-infected cells during the early stages of infection.AimTo test the hypothesis that circulating T cells exhibit phenotypic and functional activation characteristics during the viremic phase of ZIKV infection.MethodsA comprehensive analysis using mass cytometry was performed on peripheral blood mononuclear cells obtained from patients with acute ZIKV infection (as confirmed by RT-PCR) and compared with that from healthy donors (HD). The frequency of IFN-γ-producing T cells in response to peptide pools covering immunogenic regions of structural and nonstructural ZIKV proteins was quantified using an ELISpot assay.ResultsCirculating CD4+ and CD8+ T lymphocytes from ZIKV-infected patients expressed higher levels of IFN-γ and pSTAT-5, as well as cell surface markers associated with proliferation (Ki-67), activation ((HLA-DR, CD38) or exhaustion (PD1 and CTLA-4), compared to those from HD. Activation of CD4+ and CD8+ memory T cell subsets, including Transitional Memory T Cells (TTM), Effector Memory T cells (TEM), and Effector Memory T cells Re-expressing CD45RA (TEMRA), was prominent among CD4+ T cell subset of ZIKV-infected patients and was associated with increased levels of IFN-γ, pSTAT-5, Ki-67, CTLA-4, and PD1, as compared to HD. Additionally, approximately 30% of ZIKV-infected patients exhibited a T cell response primarily directed against the ZIKV NS5 protein.ConclusionCirculating T lymphocytes spontaneously produce IFN-γ and express elevated levels of pSTAT-5 during the early phase of ZIKV infection whereas recognition of ZIKV antigen results in the generation of virus-specific IFN-γ-producing T cells.

Abstract Background Fibrosis is defined as an excessive accumulation of extracellular matrix (ECM) components. Many organs are subjected to fibrosis including the lung, liver, heart, skin, kidney, and muscle. Muscle fibrosis occurs in response to trauma, aging, or dystrophies and impairs muscle function. Fibrosis represents a hurdle for the treatment of human muscular dystrophies. While data on the mechanisms of fibrosis have mostly been investigated in mice, dystrophic mouse models often do not recapitulate fibrosis as observed in human patients. Consequently, the cellular and molecular mechanisms that lead to fibrosis in human muscle still need to be identified. Methods Combining mass cytometry, transcriptome profiling, in vitro co‐culture experiments, and in vivo transplantation in immunodeficient mice, we investigated the role and nature of nonmyogenic cells (fibroadipogenic progenitors, FAPs) from human fibrotic muscles of healthy individuals (FibMCT) and individuals with oculopharyngeal muscular dystrophy (OPMD; FibMOP), as compared with nonmyogenic cells from human nonfibrotic muscle (MCT). Results We found that the proliferation rate of FAPs from fibrotic muscle is 3–4 times higher than those of FAPs from nonfibrotic muscle (population doubling per day: MCT 0.2 ± 0.1, FibMCT 0.7 ± 0.1, and FibMOP 0.8 ± 0.3). When cocultured with muscle cells, FAPs from fibrotic muscle impair the fusion index unlike MCT FAPs (myoblasts alone 57.3 ± 11.1%, coculture with MCT 43.1 ± 8.9%, with FibMCT 31.7 ± 8.2%, and with FibMOP 36.06 ± 10.29%). We also observed an increased proliferation of FAPs from fibrotic muscles in these co‐cultures in differentiation conditions (FibMCT +17.4%, P

Myasthenia Gravis (MG) is a neurological autoimmune disease characterized by disabling muscle weaknesses due to anti-acetylcholine receptor (AChR) autoantibodies. To gain insight into immune dysregulation underlying early-onset AChR+ MG, we performed an in-depth analysis of peripheral mononuclear blood cells (PBMCs) using mass cytometry. PBMCs from 24 AChR+ MG patients without thymoma and 16 controls were stained with a panel of 37 antibodies. Using both unsupervised and supervised approaches, we observed a decrease in monocytes, for all subpopulations: classical, intermediate, and non-classical monocytes. In contrast, an increase in innate lymphoid cells 2 (ILC2s) and CD27- γδ T cells was observed. We further investigated the dysregulations affecting monocytes and γδ T cells in MG. We analyzed CD27- γδ T cells in PBMCs and thymic cells from AChR+ MG patients. We detected the increase in CD27- γδ T cells in thymic cells of MG patients suggesting that the inflammatory thymic environment might affect γδ T cell differentiation. To better understand changes that might affect monocytes, we analyzed RNA sequencing data from CD14+ PBMCs and showed a global decrease activity of monocytes in MG patients. Next, by flow cytometry, we especially confirmed the decrease affecting non-classical monocytes. In MG, as for other B-cell mediated autoimmune diseases, dysregulations are well known for adaptive immune cells, such as B and T cells. Here, using single-cell mass cytometry, we unraveled unexpected dysregulations for innate immune cells. If these cells are known to be crucial for host defense, our results demonstrated that they could also be involved in autoimmunity.

Efforts to develop effective strategies that improve dietary intake are needed; however, this improvement in diet quality must not be at the expense of well-being. The Well-Being related to Food Questionnaire (Well-BFQ©) is a tool that has been developed in France to comprehensively measure food well-being. Even though the same language is spoken in France and in Québec, cultural and linguistic differences are present, which supports the importance of adapting and validating this tool before its use in the Québec population. This study aimed to adapt and validate the Well-BFQ© for the French-speaking general adult population of Québec, Canada. The Well-BFQ© underwent a full linguistic adaptation process, including an expert panel adaptation step, a pretest among 30 French-speaking adult (18–65 years) Quebecers, and a final proofreading. The questionnaire was thereafter administered to 203 French-speaking adult Quebecers (49.3% females, MAGE = 34.9, SD = 13.5; 88.2% Caucasians; 54.2% with a university degree). The exploratory factor analysis showed a two-factor structure: (1) food well-being related to physical and psychological health (27 items) and (2) food well-being related to symbolic/pleasure of food (32 items). Internal consistency was adequate, with a Cronbach’s α of 0.92 and 0.93, respectively, for the subscales, and 0.94 for the total scale. The total food well-being score, as well as the two subscale scores, were associated with psychological and eating-related variables in expected directions. Overall, the adapted version of the Well-BFQ© was found to be a valid instrument to measure food well-being in the French-speaking general adult population of Québec, Canada.

Fabry disease (FD) is an X‐linked disease characterized by an accumulation of glycosphingolipids, notably of globotriaosylceramide (Gb3) and globotriaosylsphingosine (lysoGb3) leading to renal failure, cardiomyopathy, and cerebral strokes. Inflammatory processes are involved in the pathophysiology. We investigated the immunological phenotype of peripheral blood mononuclear cells in Fabry patients depending on the clinical phenotype, treatment, Gb3, and lysoGb3 levels and the presence of anti‐drug antibodies (ADA). Leucocytes from 41 male patients and 20 controls were analyzed with mass cytometry using both unsupervised and supervised algorithms. FD patients had an increased expression of CD27 and CD28 in memory CD45‐ and CD45 + CCR7‐CD4 T cells (respectively p < 0.014 and p < 0.02). Percentage of CD45RA‐CCR7‐CD27 + CD28+ cells in CD4 T cells was correlated with plasma lysoGb3 (r = 0.60; p = 0.0036) and phenotype (p < 0.003). The correlation between Gb3 and CD27 in CD4 T cells almost reached significance (r = 0.33; p = 0.058). There was no immune profile associated with the presence of ADA. Treatment with agalsidase beta was associated with an increased proportion of Natural Killer cells. These findings provide valuable insights for understanding FD, linking Gb3 accumulation to inflammation, and proposing new prognostic biomarkers. [ABSTRACT FROM AUTHOR]

The importance of the adaptive T cell response in the control and resolution of viral infection has been well established. However, the nature of T cell–mediated viral control mechanisms in life-threatening stages of COVID-19 has yet to be determined. The aim of the present study was to determine the function and phenotype of T cell populations associated with survival or death of patients with COVID-19 in intensive care as a result of phenotypic and functional profiling by mass cytometry. Increased frequencies of circulating, polyfunctional CD4+CXCR5+HLA-DR+ stem cell memory T cells (Tscms) and decreased proportions of granzyme B–expressing and perforin-expressing effector memory T cells were detected in recovered and deceased patients, respectively. The higher abundance of polyfunctional PD-L1+CXCR3+CD8+ effector T cells (Teffs), CXCR5+HLA-DR+ Tscms, and anti-nucleocapsid (anti-NC) cytokine-producing T cells permitted us to differentiate between recovered and deceased patients. The results from a principal component analysis show an imbalance in the T cell compartment that allowed for the separation of recovered and deceased patients. The paucity of circulating PD-L1+CXCR3+CD8+ Teffs and NC-specific CD8+ T cells accurately forecasts fatal disease outcome. This study provides insight into the nature of the T cell populations involved in the control of COVID-19 and therefore might impact T cell–based vaccine designs for this infectious disease.

Protozoan parasites in food or water samples are generally detected using microscopy or PCR followed by Sanger sequencing. However, microscopy is subjective, requires a high degree of expertise and has limited sensitivity, while DNA sequencing requires expensive and specialized equipment and facilities. This study describes a cloth-based hybridization array system (CHAS) that is an alternative to Sanger sequencing to confirm PCR-positive samples. CHAS is an inexpensive, rapid and reliable method for the simultaneous detection of multiple protozoan parasite species based on the colorimetric detection of PCR amplicons on a polyester cloth. PCR primers and CHAS hybridization probes were developed to detect the protozoan parasites Giardia duodenalis, Cryptosporidium spp. and Toxoplasma gondii. In addition, CHAS probes were designed for the differentiation of G. duodenalis Assemblages A and B. In artificially contaminated fresh produce (lettuce, parsley) and water samples (river water, wastewater), this CHAS assay allowed for the successful detection of G. duodenalis, Cryptosporidium spp., and T. gondii. The present study demonstrates that the CHAS detection method is a simple and inexpensive alternative to DNA sequencing for the confirmation of PCR-positive results in laboratories testing for parasites in food or water samples. This assay may also be beneficial in developing countries, where DNA sequencing facilities may not be readily available.

The Aid and Attendance (A&A) benefit is a cash entitlement for Veterans who served in the U.S. military to obtain personal care services. Our objective was to identify factors contributing to variation in A&A enrollment across VA Medical Centers (VAMCs). We used VA data to calculate the enrollment rate among older Veterans receiving a VA pension or compensation in 2015, then purposefully sampled social work leaders at 15 VAMCs with the highest (n = 7) and lowest (n = 8) enrollment rates for interviews. All respondents viewed A&A as an important benefit. Participants at high-enrollment sites indicated strong working relationships with Veterans Benefits Administration (VBA) and Veterans Service Organizations (VSOs) with onsite presence and education about A&A facilitate access. Participants at low-enrollment sites indicated they desired education around A&A eligibility criteria and collaboration with VBA/VSOs. VA and non-VA social workers would benefit from education about VBA's benefits, and this requires collaboration with VBA representatives. [ABSTRACT FROM AUTHOR]

Children today are spending less time than ever outdoors, contributing to a culture of environmental apathy and separation from the natural world. In the growing field of environmental education, teachers are challenged to introduce the outdoors into their curriculum. In Vermont, some public school teachers have successfully implemented environmental education into their practice in a variety of interesting ways. This paper analyzes a series of teacher interviews and observations in Vermont public schools to identify successful strategies and pedagogies being utilized for environmental education. Several common themes emerged from this study, including place-based education, hands-on exploration, and free choice learning.