Journal: Stem Cell Reports

Article Title: Deciphering the differential impact of thrombopoietin/MPL signaling on hematopoietic stem/progenitor cell function in bone marrow and spleen

doi: 10.1016/j.stemcr.2023.12.004

Figure Lengend Snippet: The marrow and splenic hematopoietic niche in TPO −/− and MPL −/− mice (A) Representative whole-mount confocal image of thick femur sections, in which the vasculature was stained intravenously with anti-VE-cadherin antibody (20× magnification, three 5-μm Z -stacks per image; scale bar, 100 μm). Quantification of VE-cadherin + vasculature (white) area in the marrow of young control, TPO −/− , and MPL −/− mice (n = 2–3 mice in each group). A total of 12–16 non-overlapping 450 × 350 pixel areas at 20× magnification were analyzed for each group. (B) Representative image of thin spleen sections in which the vasculature was stained intravenously with anti-VE-cadherin antibody (4× magnification; scale bar, 500 μm). Quantification of VE-cadherin + vasculature (purple) area in the spleen of young control, TPO −/− , and MPL −/− mice (n = 2–3 mice in each group). There were 16 non-overlapping areas (normalized to 450 × 350 pixels) at 4× magnification that were analyzed for each group. (C–E) Representative flow cytometry plot of CD45 − CD31 + ECs (C) and flow cytometry quantitative analysis of membranal SCF (D) and intracellular CXCL12 (E) expression in marrow ECs and spleen ECs of young control (n = 3–6), TPO −/− (n = 3–6), and MPL −/− (n = 3–6) mice. MFI, median fluorescence intensity. (F) Representative flow cytometry plots showing the gating strategy of CD45 − CD31 – Ter119 – Sca1 – perivascular stromal cells. (G) Flow cytometry quantitative measurement of perivascular stromal cells in the marrow (left) and spleen (right) of young control (n = 3), TPO −/− (n = 3), and MPL −/− (n = 3) mice. (H) CXCL12 expression in marrow (left) and spleen (right) perivascular stromal cells of young control (n = 3), TPO −/− (n = 3), and MPL −/− (n = 3) mice measured using flow cytometry.

Article Snippet: Intracellular CXCL12 expression and membranal SCF expression were analyzed using an anti-mouse CXCL12 antibody (Clone 79018, R&D Systems, Minneapolis, MN, USA) and an anti-mouse SCF antibody (Clone #40215, R&D Systems) as we previously described ( ).

Techniques: Staining, Flow Cytometry, Expressing, Fluorescence

Journal: Cell death & disease

Article Title: SCF and IL-33 regulate mouse mast cell phenotypic and functional plasticity supporting a pro-inflammatory microenvironment.

doi: 10.1038/s41419-023-06139-7

Figure Lengend Snippet: Fig. 7 SCF controls connective tissue-like MC accumulation in tumor lesions. A Schematic representation of anti-SCF neutralizing Ab administration in vivo. AOM/DSS-treated mice were i.p. injected three times with anti-SCF or control Ab (100 μg/mouse) starting from the fourth DSS cycle. Treated and control mice were sacrificed at 13 weeks from AOM administration. B Colon paraffin-embedded sections from AOM/DSS mice treated with Ctrl-Ig or anti-SCF neutralizing antibodies as described in (A) were stained with anti-MCP4 Ab followed by Alexa Fluor 488 secondary Abs (green). Nuclei were counterstained with DAPI (blue) and images were acquired with a Zeiss LSM980 confocal microscopy using a 20× objective. The frequencies of MCs positive for mMCP4 protease were analyzed in 20 fields randomly acquired from tumor lesions and shown as mean ± SD cells/field. Paired Student’s t test: *p < 0.05. Number of adenomas and colon lengths are shown in (C). Paired Student’s t test: *p < 0.05. Graphs are representative of two independent experiments with a total of 5 mice/group.

Article Snippet: For blocking experiments, mice were i.p. injected with anti-SCF neutralizing antibody (AB-455-NA R&D Systems) or normal goat control IgG (AB-108-C R&D Systems) (100μg/mouse) three times every 10 days starting one day before the fourth DSS cycle (see Fig. 7A).

Techniques: In Vivo, Injection, Control, Staining, Confocal Microscopy