1. |
- Beal, Jacob, et al.
(författare)
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Robust estimation of bacterial cell count from optical density
- 2020
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Ingår i: Communications Biology. - : Springer Science and Business Media LLC. - 2399-3642. ; 3:1
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Tidskriftsartikel (refereegranskat)abstract
- Optical density (OD) is widely used to estimate the density of cells in liquid culture, but cannot be compared between instruments without a standardized calibration protocol and is challenging to relate to actual cell count. We address this with an interlaboratory study comparing three simple, low-cost, and highly accessible OD calibration protocols across 244 laboratories, applied to eight strains of constitutive GFP-expressing E. coli. Based on our results, we recommend calibrating OD to estimated cell count using serial dilution of silica microspheres, which produces highly precise calibration (95.5% of residuals <1.2-fold), is easily assessed for quality control, also assesses instrument effective linear range, and can be combined with fluorescence calibration to obtain units of Molecules of Equivalent Fluorescein (MEFL) per cell, allowing direct comparison and data fusion with flow cytometry measurements: in our study, fluorescence per cell measurements showed only a 1.07-fold mean difference between plate reader and flow cytometry data.
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2. |
- Borgström, Emilie W., et al.
(författare)
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Three Adult Cases of STAT1 Gain-of-Function with Chronic Mucocutaneous Candidiasis Treated with JAK Inhibitors
- 2023
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Ingår i: Journal of Clinical Immunology. - : Springer. - 0271-9142 .- 1573-2592. ; 43, s. 136-150
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Tidskriftsartikel (refereegranskat)abstract
- Purpose; The aim of this study was to characterize clinical effects and biomarkers in three patients with chronic mucocutaneous candidiasis (CMC) caused by gain-of-function (GOF) mutations in the STAT1 gene during treatment with Janus kinase (JAK) inhibitors.Methods: Mass cytometry (CyTOF) was used to characterize mononuclear leukocyte populations and Olink assay to quantify 265 plasma proteins. Flow-cytometric Assay for Specific Cell-mediated Immune-response in Activated whole blood (FASCIA) was used to quantify the reactivity against Candida albicans.Results: Overall, JAK inhibitors improved clinical symptoms of CMC, but caused side effects in two patients. Absolute numbers of neutrophils, T cells, B cells, and NK cells were sustained during baricitinib treatment. Detailed analysis of cellular subsets, using CyTOF, revealed increased expression of CD45, CD52, and CD99 in NK cells, reflecting a more functional phenotype. Conversely, monocytes and eosinophils downregulated CD16, consistent with reduced inflammation. Moreover, T and B cells showed increased expression of activation markers during treatment. In one patient with a remarkable clinical effect of baricitinib treatment, the immune response to C. albicans increased after 7 weeks of treatment. Alterations in plasma biomarkers involved downregulation of cellular markers CXCL10, annexin A1, granzyme B, granzyme H, and oncostatin M, whereas FGF21 was the only upregulated marker after 7 weeks. After 3 months, IFN-gamma and CXCL10 were downregulated.Conclusions: The clinical effect of JAK inhibitor treatment of CMC is promising. Several biological variables were altered during baricitinib treatment demonstrating that lymphocytes, NK cells, monocytes, and eosinophils were affected. In parallel, cellular reactivity against C. albicans was enhanced.
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3. |
- Consiglio, Camila, et al.
(författare)
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Immune system adaptation during gender-affirming testosterone treatment
- 2023
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Ingår i: Journal of Reproductive Immunology. - : Elsevier. - 0165-0378 .- 1872-7603. ; 159, s. 29-30
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Tidskriftsartikel (övrigt vetenskapligt/konstnärligt)abstract
- Biological sex impacts human immune responses, modulating susceptibility and severity to immune-related diseases. Female generally mount more robust immune responses than males, resulting in lower infection severity and greater autoimmunity incidence. Here, we addressed the contribution of testosterone to human immune function by analyzing a cohort of subjects undergoing gender-affirming testosterone treatment. We performed systems-level immunomonitoring through mass cytometry, scRNA and scA-TAC-Sequencing, and proteome profiling of blood samples at baseline and following 3 and 12 months of treatment. Testosterone treatment was associated with a low-grade inflammatory profile, evidenced by upregulation of proinflammatory plasma proteome (e.g., EN-RAGE, OSM, TNF), and induction of an inflammatory transcriptional program associated with NFkB signaling, and TNF signaling. Following testosterone treatment, higher NFkB activity was revealed in CD4 T, CD8 T, and NK cells in scATACseq analyses. Further, testosterone increased monocytic inflammatory responses upon bacterial stimulation in vitro. Although testosterone was associated with this inflammatory profile, it also exerted negative effects on antiviral immunity. Firstly, the percentage of plasmacytoid dendritic cells (pDC) decreased over transition, with pDC also displaying phenotypic changes associated with lower IFN responses. Secondly, bulk transcriptomics analyses show an overall reduction of IFNa responses. Thirdly, testosterone treatment led to reduced IFNa production upon PBMCs stimulation with a viral agonist. Our results show that testosterone has broad effects on the human immune system, and significantly modulates important players in antiviral immunity and inflammatory response. Identifying pathways involved in immune sexual dimorphism will help define novel targets for effective prevention and treatment of immune-mediated diseases.
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4. |
- Consiglio, Camila Rosat, et al.
(författare)
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The Immunology of Multisystem Inflammatory Syndrome in Children with COVID-19
- 2020
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Ingår i: Cell. - : CELL PRESS. - 0092-8674 .- 1097-4172. ; 183:4, s. 968-981
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Tidskriftsartikel (refereegranskat)abstract
- Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is typically very mild and often asymptomatic in children. A complication is the rare multisystem inflammatory syndrome in children (MIS-C) associated with COVID-19, presenting 4-6 weeks after infection as high fever, organ dysfunction, and strongly elevated markers of inflammation. The pathogenesis is unclear but has overlapping features with Kawasaki disease suggestive of vasculitis and a likely autoimmune etiology. Weapply systems-level analyses of blood immune cells, cytokines, and autoantibodies in healthy children, children with Kawasaki disease enrolled prior to COVID-19, children infected with SARS-CoV-2, and children presenting with MIS-C. We find that the inflammatory response in MIS-C differs from the cytokine storm of severe acute COVID-19, shares several features with Kawasaki disease, but also differs from this condition with respect to T cell subsets, interleukin (IL)-17A, and biomarkers associated with arterial damage. Finally, autoantibody profiling suggests multiple autoantibodies that could be involved in the pathogenesis of MIS-C.
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