Journal: bioRxiv

Article Title: A Novel Humanized Mouse Model for HIV and Tuberculosis Co-infection Studies

doi: 10.1101/2024.03.05.583545

Figure Lengend Snippet: (A) Lung sections were obtained from formalin-fixed tissues of animals in all experimental groups (one animal for each group) and subjected to hematoxylin-eosin staining, two different amplifications are shown. (B) Immunofluorescence staining of surface markers for human macrophages (CD68-Alexafluor 568, in orange) and B-cells (CD19-Alexa 488, in green) in lung sections from uninfected and Mtb -infected mice. DAPI-supplemented mounting buffer (in blue) was used for nuclei staining. (C) Representative 3D renditions of CT scan and lung volume pictures obtained from animals in all experimental groups. (D) Pulmonary function test parameters: Resistance (Rrs), compliance (Crs) and elastance (Ers), were collected from animals in all experimental groups at the end of the trial (Uninfected: n=5; HIV-infected: n=8; Mtb -infected: n=8; HIV/ Mtb -co-infected: n=7).

Article Snippet: Primary antibody incubation was carried out overnight at 4 °C with human-CD68 monoclonal antibody (cat. No. 14-0688-82, Invitrogen) and CD19 Rabbit polyclonal antibody (cat. No. 27949-1-AP, Proteintech, Rosemont, USA), diluted in PBS + 0.4% triton + 1% FBS at the recommended dilutions.

Techniques: Staining, Immunofluorescence, Infection, Computed Tomography

Journal: medRxiv

Article Title: Longitudinal multi-omics analysis identifies responses of megakaryocytes, erythroid cells and plasmablasts as hallmarks of severe COVID-19 trajectories

doi: 10.1101/2020.09.11.20187369

Figure Lengend Snippet: a , Schematic workflow of the scRNA-seq analyses performed on the B cell compartment identified in . b , B cell compartment subtypes represented as a UMAP. In total, 22,190 cells are depicted. Clusters represented are Memory B cells (MB, dark red), naïve B cells (N, red), transitional B cells (trans-orange) and plasmablast (PB, blue). c , B cell compartment pseudotimes represented as a UMAP. d , Flow cytometry analysis of B cell subtypes. CD19 + B cells were stained for CD20 and CD27. CD20 - CD27 high B cells classified as PB, CD20 + CD27 + cells as MB and CD20 + CD27 - cells as N. Proportions of each cell type among CD19 + B cells were plotted against the sampling time relative to the disease onset. The points were coloured by corresponding pseudotime and connected by patient. Only patients from Kiel cohort ( n = 7 individuals) are shown. e , Plasmablast-specific UMAP highlighted neutrophil-like cells (NL). Smaller UMAPs corresponding to expression of plasmablast markers ( CD27 , CD38 , and TNFRSF17 ) and neutrophil-like markers ( ELANE , MPO , and CAMP ). f , Cell trajectory analysis of B cell compartment. Cell trajectory was calculated using Monocle3 and B cell subtypes are displayed in left panel followed by the corresponding transition-pseudotime analysis rooted on transitional B cells (purple) and differentiated into 2 branches: B cells naïve/memory branch (grey line/culminating in yellow) and an over imposed plasmablast branch, from which most of the plasmablasts (black line/culminating in orange) are not connected to neutrophil-like cells (grey). g , Dot plot for pseudotime signature genes in plasmablasts. Genes were selected based on ten most characteristic upregulated genes. Colour discriminates upregulated (red) or downregulated (blue) genes, while point sizes represents the number of cells per group expressing the corresponding gene. h , GO enrichment analysis for upregulated genes during disease trajectory. Size of the dots is proportional to the gene ratio and the colour corresponds to the p -value of the enrichment. Selected top terms are visualized. i , Gene expression of genes of interest in B cell subtypes, pseudotimes and cohort 2. Genes of interest selected based on their high expression in PB or NL and upregulated during disease ( DERL3 , CD38, PIM2, IFI6 , XBP1 , and SLC1A4 ). For each gene, top violin plot depicting B cell subtypes expression, centre violin plot based on pseudotime expressions and bottom violin plot based on the expression of cohort 2 divided by disease classification, healthy control (white), mild disease (light grey) and severe disease (dark grey). j , Metabolic pathways enriched in B cell compartment subtypes. Top 20 active metabolic pathways for context-specific metabolic networks reconstructed in plasmablasts are shown. For each B cell subtype, significant differences in metabolic activity were determined using a Kruskal-Wallis test. Number of reaction counts found per pathways is displayed as colour intensity.

Article Snippet: B cell subsets were stained using antibodies against CD19 (clone REA675, Miltenyi), CD20 (clone REA780, Miltenyi), CD27 (clone M-T271, Biolegend, San Diego, California, US), CD138 (clone BB4/MI15, Biolegend), HLA-DR (clone REA332, Miltenyi), IgD (clone REA740, Miltenyi), IgM (clone MHM-88, Biolegend), IgA (clone M24A, Merck Millipore), CD95 (clone DX2, Biolegend) as well as CD3 (clone OKT3, Biolegend) and CD14 (clone M5E2, Biolegend) for the dump channel.

Techniques: Flow Cytometry, Staining, Sampling, Expressing, Gene Expression, Control, Activity Assay