Journal: Cell

Article Title: SARS-CoV-2 infection triggers profibrotic macrophage responses and lung fibrosis

doi: 10.1016/j.cell.2021.11.033

Figure Lengend Snippet: CD163 + macrophages accumulate in the lung in severe COVID-19 (A) Overview of study design and analyses. CT, computed tomography; BAL, bronchoalveolar lavage; scRNA-seq, single-cell RNA sequencing; snRNA-seq, single-nucleus RNA sequencing; IHC, immunohistochemistry; IF, immunofluorescence microscopy; MELC, multi-epitope ligand cartography; EM, electron microscopy; VCin, inspiratory vital capacity; PBMC, peripheral blood mononuclear cells; IAV, Influenza A virus. (B) Postmortem analysis of consecutive histological sections of non-COVID-19 (left) and COVID-19 autopsy lung samples (right) by hematoxylin and eosin (H&E; top) and CD68 IHC (bottom). Scale bar, 100 μm. (C) IF of CD68 (green) and CD163 (red) in lung tissue autopsy samples of COVID-19 patients and non-COVID-19 controls. Arrows indicate CD68 + CD163 – macrophages, and arrowheads indicate CD68 + CD163 + macrophages. Scale bar, 20 μm. (D) Quantification of CD68 + macrophage density (left) and the proportion of CD163 + macrophages (right) in lung autopsy samples from fifteen donors (as in C). Mann-Whitney test; ∗ p < 0.05. (E) Representative images of consecutive histological sections of lung autopsy samples. H&E (left), CD68 IHC (middle), and SARS-CoV-2 RNA-FISH (right). Arrowheads indicate SARS-CoV-2 RNA-positive macrophages. Scale bars, 50 μm, 25 μm. RNA-FISH, RNA-fluorescence in situ hybridization. (F) Lung autopsy samples of 9 COVID-19 patients were analyzed by MELC with a panel of 22 markers on 19 fields of view (FOVs). Two-dimensional embedding computed by UMAP on 9,684 computationally identified CD45 positive cells (T cells, CD3 + ; B cells, CD20 + ; NK cells, CD56 + ; neutrophils, MRP14 + /CD66b + ; monocytes, MRP14 + /CCR2 + ; macrophages, MRP14 + /HLA-DR + ). (G) Relative proportion (of total CD45 + cells) of cell types in all 19 FOVs (left), and average cell numbers (summary, right).

Article Snippet: CD66b-PE , Miltenyi Biotec , Cat# 130-122-922, N/A.

Techniques: Computed Tomography, RNA Sequencing Assay, Immunohistochemistry, Immunofluorescence, Microscopy, Electron Microscopy, MANN-WHITNEY, Fluorescence, In Situ Hybridization

Journal: Cell

Article Title: SARS-CoV-2 infection triggers profibrotic macrophage responses and lung fibrosis

doi: 10.1016/j.cell.2021.11.033

Figure Lengend Snippet:

Article Snippet: CD66b-PE , Miltenyi Biotec , Cat# 130-122-922, N/A.

Techniques: Immunohistochemistry, Plasmid Preparation, Recombinant, Staining, Lysis, Protease Inhibitor, Mass Spectrometry, Sequencing, Modification, Bicinchoninic Acid Protein Assay, Enzyme-linked Immunosorbent Assay, Software

Journal: bioRxiv

Article Title: Beta-2-microglobulin stimulates neutrophil phagocytosis of bacteria and apoptotic cells

doi: 10.1101/2025.10.30.685267

Figure Lengend Snippet: Efferocytosis of apoptotic Jurkat cells labeled with CellTrace™ Violet by CD66b-positive granulocytes stained with CellTrace™ Far Red was examined under various conditions. (A) PMNs were exposed to varying concentrations of β2m or dK58β2m in the presence of apoptotic Jurkat cells. (B) PMNs were treated with apoptotic cells alone or in combination with 50 µg/ml β2m, 50 µg/ml dK58β2m, 100 ng/ml GM-CSF, or combinations thereof. (C-D) PMNs were treated with apoptotic Jurkat cells with 50 µg/ml β2m (C) or dK58β2m (D) combined with 10 or 25% human AB serum. In all experiments, a 4:1 apoptotic cell-to-PMN ratio was used. Treatment with 100 ng/ml GM-CSF served as a positive control, and pre-incubation with 10 µg/ml cytochalasin D (cyto D) served as a negative control. Results are shown as mean ± SD of double-positive PMNs. Sample sizes were n=6 or n=4 (A), n=10 (B), and n=8 or 3 donors (C and D). Group comparisons were analyzed using mixed-effects analysis and Tukey’s multiple comparisons test (A, C, and D) or repeated-measures ANOVA and Tukey’s multiplecomparisons test(B). Statistical significance is indicated by *P < 0.05 and **P < 0.01, ***P < 0.001, ****P < 0.0001.

Article Snippet: As a control for phagocytic uptake, PMNs were pre-treated with 10 μg/ml cytochalasin D (Thermo Fisher Scientific, #PHZ1063) for 20 min. After the two-hour incubation, the cells were washed in HBSS, and granulocytes were stained with anti-CD66b antibodies (R&D systems, #FAB4246G) for 30 min at 4°C in the dark.

Techniques: Labeling, Staining, Positive Control, Incubation, Negative Control

Journal: bioRxiv

Article Title: Immune Checkpoint Molecules as Biomarkers of Staphylococcus aureus Bone Infection and Clinical Outcome

doi: 10.1101/2024.12.30.630837

Figure Lengend Snippet: Bone tissues surgically removed from PJI patient with S. aureus osteomyelitis were processed for histology and immunohistochemistry. (A) Representative 100x image (bar =100 μm) of a H&E-stained section is shown to illustrate the inflammatory cells within the region of interest (box). ( B-D) Parallel histology sections containing the region of interest were immunostained with labelled antibodies against CD3, PD1, S. aureus, TIM-3 (green), LAG-3, and CD66b, counter stained with DAPI, and representative fluorescent microscopy images are shown at 200x (bar = 100 μm). ( B ) Note CD3 + /PD1 + T cells detected in areas of S. aureus infection (white arrows). (C) Note CD3 + /TIM-3 + (white arrows) and CD3 + /LAG-3 + (yellow arrows) T cells at the site of S. aureus infection. (D) Note TIM-3 + /CD66b+ neutrophils at the site of infection (white arrows).

Article Snippet: Primary antibodies: The following antibodies were utilized for immunostaining: goat anti-CD3ε (clone M-20, sc-1127, RRID:AB_631128, Santa Cruz Biotechnology), mouse anti-PD-1 (10377-MM23, RRID:AB_2936309, Sino Biologicals), Rabbit anti-LAG3 (clone BLR027F, NBP2-76402, RRID:AB_3403543, Novus Biologicals), Mouse anti-TIM3/HAVCR2 (clone TIM3/4031, V8754-20UG, NSJ Bioreagents), Rabbit anti- S. aureus (PA1-7246, RRID:AB_561546, Thermo Fisher Scientific), and Mouse anti-CD66b (G10F5, NBP2-80664, RRID:AB_3096017, Novus Biologicals).

Techniques: Immunohistochemistry, Staining, Microscopy, Infection

Journal: JBMR Plus

Article Title: Staphylococcus aureus Panton-Valentine Leukocidin worsens acute implant-associated osteomyelitis in humanized BRGSF mice

doi: 10.1093/jbmrpl/ziad005

Figure Lengend Snippet: HuBRGSF mice infected with isogenic PVL mutant had fewer human myeloid cells and dead cells in the bone niche. Myeloid cell populations in bone marrow of huBRGSF mice with a transtibial implant-associated osteomyelitis after infection with MRSA USA300 WT, USA300 Δpvl, or USA300 Δpvl+pvl. Percentages of (A) human CD45 + myeloid cells, (B) neutrophils, (C) monocytes/macrophages, (D) dendritic cells, (E) NK cells, (F) HLA-DR + neutrophils, (G) HLA-DR + monocytes/macrophages, or (H) eFluor780 + dead cells in bone marrow of WT- (circle), Δpvl- (square), or Δpvl+pvl- (triangle) infected mice, 3 d after surgery are depicted. Data are mean ± SD and was analyzed with a Tukey’s multiple comparison test of a one-way ANOVA. N = 6 for WT-infected mice, n = 5 for Δpvl-infected mice, and n = 6 for Δpvl+pvl-infected mice. * p < 0.05, * * p < 0.01, * * * p < 0.001.

Article Snippet: The following primary antibodies were used for immunofluorescent staining of HuBRGSF infected limbs (USA300 WT, USA300 Δpvl, or USA300 Δpvl+pvl): mouse anti-CEACAM8/CD66b for neutrophils (Clone G10F5, NBP2–80664, Novus Biologicals, Zug, Switzerland), rabbit anti- S. aureus (PA1–7246, RRID:AB_561546, Thermo Fisher Scientific), and mouse anti-CD68 for monocytes (Clone PG-M1, MS-1808-S1, RRID:AB_149350, Thermo Fisher Scientific).

Techniques: Infection, Mutagenesis, Comparison