Journal: Nature Communications

Article Title: Autoantibody landscape and functional role of anti-C-C motif chemokine receptor 8 autoantibodies in systemic sclerosis: post-hoc analysis of a B-cell depletion trial

doi: 10.1038/s41467-025-66974-4

Figure Lengend Snippet: A Representative histograms from flow cytometry showing binding of anti-C-C motif chemokine receptor 8 (CCR8)-positive IgG from systemic sclerosis (SSc) patients or healthy controls (HC) to CCR8-overexpressing HEK293 cells. A fluorophore-conjugated anti-human IgG Fc antibody was used for detection. B Quantification of mean fluorescence intensity (MFI) in CCR8-overexpressing cells incubated with anti-CCR8-positive or control IgG, with or without preincubation with blocking anti-CCR8 monoclonal antibody ( n = 4 technical replicates per group). C ERK phosphorylation levels in CCR8-overexpressing HEK293 cells after stimulation with CCL1 in the presence of anti-CCR8-positive or control IgG, as measured by ELISA ( n = 8 biological replicates per group). D Treg migration assay in Transwell culture systems. The number of migrated Tregs in response to CCL1 with anti-CCR8-positive IgG or control IgG ( n = 6 biological replicates per group) are presented. Statistical significance was assessed using Welch’s ANOVA with Dunnett’s T3 multiple comparison adjustment. Error bars are defined as the standard error of the mean. Source data are provided as a Source Data file.

Article Snippet: Tregs were isolated from human peripheral blood using the MACSxpress Whole Blood Treg Isolation Kit (#130-109-557, Miltenyi Biotec, Bergisch Gladbach, North Rhine-Westphalia, Germany), following the manufacturer’s instructions with minor modifications.

Techniques: Flow Cytometry, Binding Assay, Fluorescence, Incubation, Control, Blocking Assay, Phospho-proteomics, Enzyme-linked Immunosorbent Assay, Migration, Comparison

Journal: Nature Communications

Article Title: Autoantibody landscape and functional role of anti-C-C motif chemokine receptor 8 autoantibodies in systemic sclerosis: post-hoc analysis of a B-cell depletion trial

doi: 10.1038/s41467-025-66974-4

Figure Lengend Snippet: A Experimental protocol of the skin sclerosis mouse model. Six-week-old female C57BL/6NcrSlc mice (MGI ID: MGI:5295404) received daily subcutaneous injections of bleomycin (BLM, 200 µg) or PBS as a control for two weeks. Mice were also treated intraperitoneally with either control (Ctrl) IgG or anti-C-C motif chemokine receptor 8 (CCR8) antibody (Ab) once on Day 8. Skin biopsies were collected at Week 2 for histological analysis and next-generation sequencing (NGS). This figure was partially created with BioRender ( https://BioRender.com/slxff50 ). Representative histological images and quantitative analyses of dermal fibrosis and Treg infiltration in mouse skin. H&E-stained B and Masson’s trichrome–stained C skin sections show increased dermal thickness and collagen deposition following BLM treatment. Images were captured at ×200 magnification, and scale bars represent 50 μm. D Quantification of dermal thickness and E Foxp3⁺ Treg counts are presented in the accompanying bar graphs ( n = 6 biological replicates for PBS group; n = 5 biological replicates for BLM-treated groups). Statistical significance was assessed using Welch’s ANOVA with Dunnett’s T3 multiple comparison adjustment. Error bars are defined as the standard error of the mean. Source data are provided as a Source Data file.

Article Snippet: Tregs were isolated from human peripheral blood using the MACSxpress Whole Blood Treg Isolation Kit (#130-109-557, Miltenyi Biotec, Bergisch Gladbach, North Rhine-Westphalia, Germany), following the manufacturer’s instructions with minor modifications.

Techniques: Control, Next-Generation Sequencing, Staining, Comparison

Journal: bioRxiv

Article Title: UBA1 Mutations Drive RIPK1-Mediated Cell Death and Monocyte Dysfunction in VEXAS Syndrome

doi: 10.1101/2025.10.06.680650

Figure Lengend Snippet: Regulated cell death pathways are activated in myeloid cells from VEXAS patients. ( A ) Leucocytes, neutrophils, monocytes, lymphocytes count from patients with VEXAS and elderly gender-matched healthy controls (HC). Panels show individual data (dots) and means ± SEM (histograms). P values were determined by the Mann-Whitney test. ( B ) Hematoxylin-eosin staining of UBA1-mutated Sweet-like lesion revealing karyorrhectic nuclei and apoptotic debris in VEXAS (arrows). Illustrative picture is shown (Magnification x10 and x40, scale bar 100 μm and 50µm). ( C ) Representative immunofluorescence images of pMLKL (green) staining on CD14+ sorted cells form 3 active VEXAS patients and 3 elderly gender-matched HC. Nuclei are stained in blue with Hoechst. Percentage of pMLKL cell surface area per CD14+ cells, data are shown and means (Histograms) ± SEM. Forty cells were quantified for each patient. ( D ) Multiplex Immunofluorescence of skin biopsy samples from VEXAS skin lesion showing the co-expression of cleaved GSDMD, MLKL, cleaved caspase-3 and phosphorylated RIPK1 within CD68+ infiltrates in VEXAS. ( E ) Gene set enrichment analysis of apoptosis (KEGG), necroptosis (GOBP) and pyroptosis (GOBP) pathways enriched in VEXAS lesional skin (n=6) versus non lesional skin from HC (n=5) adapted form from dataset GSE245639. *P□<□0.05; **P□<□0.01; ***P□<□0.001, ****P□<□0.0001. SEM, HC, Healthy Control; Standard Error of the Mean; VEXAS, Vacuoles, E1 enzyme, X-linked, Autoinflammatory, Somatic; WT, Wild-Type.

Article Snippet: For CD14+ cells form patients, cells were sorted from 2mL fresh Whole Blood collected in EDTA tube and sorted using StraightFrom® Whole Blood CD14 MicroBeads and Whole Blood column kit (Milteny Biotec, #130-090-879) according to the manufacturer’s instructions.

Techniques: MANN-WHITNEY, Staining, Immunofluorescence, Multiplex Assay, Expressing, Control

Journal: Journal of Fungi

Article Title: Development of a Simple and Robust Whole Blood Assay with Dual Co-Stimulation to Quantify the Release of T-Cellular Signature Cytokines in Response to Aspergillus fumigatus Antigens

doi: 10.3390/jof7060462

Figure Lengend Snippet: Comparison of Aspergillus -induced cytokine release in patients with Aspergillus -associated lung pathologies and other chronic lung diseases. WB from 9 patients with Aspergillus -associated lung pathologies (2 CF with A. fumigatus sensitization, 3 ABPA, and 4 CPA) and a control group of 5 patients with other chronic lung diseases was stimulated with AfuLy or Aspf4 using the RPMI-supplemented, α-CD28-, and α-CD49d-containing stimulation tubes, as described in the Materials and Methods Section. Cytokine concentrations in plasma supernatants were quantified using a 21-plex Luminex assay. ( a , b ) Heat maps indicating median background-adjusted cytokine concentrations in the ABPA/CF/CPA patient cohort “A” and control cohort “C” depending on the antigen used for stimulation. The numeric value in the MMR column represents the median-to-median ratio between the two cohorts, with values > 1.0 indicating greater median cytokine concentrations in the ABPA/CF/CPA cohort. Δ denotes infinite median-to-median ratios (median = 0 pg/mL in the control cohort). ◊ denotes undefined median-to-median ratios (median = 0 pg/mL in both cohorts). ( c , d ) Heat maps summarizing individual cytokine response to AfuLy in all 14 patients. ( e ) Comparison of individual AfuLy-induced concentrations of selected T-helper cell signature cytokines. Medians and interquartile ranges for patients with ABPA/CF/CPA (“A”, red) and controls (“C”, grey) are indicated by black bars and colored boxes, respectively. Two-sided Mann–Whitney U test. Abbreviations: AfuLy = A. fumigatus mycelial lysate, ABPA = allergic bronchopulmonary aspergillosis, CF = cystic fibrosis, CPA = chronic pulmonary aspergillosis, Fractalk. = fractalkine, WB = whole blood.

Article Snippet: The Whole Blood Column Kit (Miltenyi Biotec, Bergisch Gladbach, Germany) was applied to a MidiMACSTM cell separator (Miltenyi Biotec, Bergisch Gladbach, Germany) and rinsed with PBS (Sigma-Aldrich, St. Louis, MO, USA) supplemented with 0.5% human AB serum (Sigma-Aldrich, St. Louis, MO, USA).

Techniques: Comparison, Control, Clinical Proteomics, Luminex, MANN-WHITNEY