bsa rat anti ly6g  (Bio X Cell)

Journal: bioRxiv

Article Title: Aspirin hastens resolution of skeletal muscle inflammation and promotes recovery of muscle strength following acute injury

doi: 10.64898/2026.04.21.719989

Figure Lengend Snippet: A: Representative histological (H & E) and immunofluorescence images of TA muscle cross-sections at day 1 post-injury following treatment with vehicle (VEH), indomethacin (INDO), aspirin (ASA), or a combination of INDO + ASA. Sections are stained for Ly6G (neutrophils, red), CD68 (total macrophages, green), CD206 (M2-like macrophages, red), with DAPI (nuclei, blue) and Laminin (LAM, white) to visualize fiber boundaries. B-E : Quantification of day 1 post-injury inflammatory markers including Ly6G + cell density ( B ), CD68 + cell density ( C ), the ratio of Ly6G + to total CD68 + cells ( D ), and the M1/M2 macrophage ratio ( E ). F: Representative H & E and immunofluorescence images at day 3 post-injury for the same treatment groups and markers. G-J: Quantitative analysis of inflammatory cell dynamics at day 3 post-injury, including Ly6G + cell density ( G ), CD68 + cell area as a percentage of total tissue ( H ), CD206 + cell densities ( I ), and the M1/M2 ratio ( J ). All data are presented as mean ± SEM. Statistical significance was determined by one-way ANOVA followed by Holm-Šídák post hoc tests. Groups labeled with different letters are significantly different from one another, while groups sharing a common letter are not significantly different.

Article Snippet: Primary antibodies used include MyHC type I [Developmental Studies Hybridoma Bank (DSHB), BA-D5c, 1:100], MyHC type IIA (DSHB, SC-71c, 1:100), MyHC type IIB (DSHB, BF-F3c, 1:100), eMHC (DSHB, F1.652s, 1:20), Ly6G (GR1) (Bio-Rad, MCA2387, 1:50), CD68 (Bio-Rad, MCA1957, 1:200), CD206 (Bio-Rad, MCA2387, 1:50), and laminin (Abcam, ab7463, 1:200).

Techniques: Immunofluorescence, Staining, Labeling

Journal: The Journal of Clinical Investigation

Article Title: Neutrophil-microglia interaction drives motor dysfunction in a neuromyelitis optica model induced by subarachnoid AQP4-IgG

doi: 10.1172/JCI199706

Figure Lengend Snippet: ( A ) Timeline for injecting neutrophil-depleting anti-Ly6G-IgG or isotype control-IgG (100 mg/kg, i.p.), inserting lumbar subarachnoid catheter, infusing AQP4-IgG, and rotarod testing. ( B ) Flow cytometry confirms neutrophil ablation efficiency (percentage CD45 + CD11b + Gr1 + MPO + cells among peripheral CD45 + CD11b + cells). ( C ) (Upper) t-SNE analysis of CD45 + immune cell subtypes from lumbar spinal cords of control and neutrophil-depleted mice. (Lower) Quantification of data in B (3 mice/group). ( D ) Representative confocal images of neutrophils in lungs of mice receiving neutrophil-depleting anti-Ly6G-IgG or isotype control-IgG (3 mice/group). ( E ) Microglial activation, reflected by Cx3cr1GFP signal, in corresponding lumbar cord regions of mice without and with neutrophil ablation (by Ly6G-IgG or isotype control-IgG) after 3 days’ infusion with normal control mouse IgG or AQP4-IgG. ( F ) Quantification of microglia-occupied areas in E ( n = 4–5 mice per group). ( G ) Motor function, reflected by rotarod test, in neutropenic mice (anti-Ly6G-IgG–treated) and non-neutrophil-ablated (isotype control-IgG–treated) during 5 days’ infusion of AQP4-IgG or normal control mouse IgG (0.1 μg/μL; time: F (2.415, 28.98) = 4.838, P = 0.0113; treatment: F (2, 12) = 12.46, P = 0.0012; interaction: F (10, 60) = 7.100, P < 0.0001; n = 5 mice per group). ( H ) Experimental design: WT mice were continuously infused with Ctrl-IgG or AQP4-IgG by osmotic pumps for 7 days from day 0; infusion was discontinued at day 8. ( I ) Motor impairment worsened progressively in AQP4-IgG recipients, with nadir at day 8. Continued rotarod testing for another 3 weeks showed progressive motor recovery from day 8. ( J ) Correlations between microglial activation state (lumbar microglial area) and latency to fall in rotarod test. Simple linear regression (1 dot represents 1 mouse at day 3 of IgG infusion). Statistics: C used t test; Tukey’s post hoc multiple comparisons test (1-way ANOVA) in F ; 2-way repeated measures ANOVA with Holm-Šídák post hoc test in G and I .

Article Snippet: For immunofluorescence staining, sections were permeabilized and blocked (30 min), using 0.25% Triton X-100 in PBS containing 2.5% bovine serum albumin and 5% donkey serum, then exposed 16–24 hours at 4°C to the following primary antibodies in PBS containing 0.1% Triton X-100 (Sigma) and 1% BSA: rat anti-Ly6G (Bio X Cell; BE0075-1, clone 1A8, 1:500), human/mouse polyclonal goat anti-MPO (R&D Systems; AF3667; 1:800), rat anti-NE (R&D Systems, MAB4517-SP, clone 887105, 1:500), mouse anti-C5 (Hycult Biotech; HM1073, clone BB5.1, 1:500), rat anti-C5a (Invitrogen; MA5-23910, clone 295108, 1:300), goat polyclonal anti-GFP (Abcam; Ab6673, 1:1,000), polyclonal goat anti-CD31 (R&D Systems; AF3628, 1:500), rabbit antiIBA1 (Abcam; Ab178846 , clone EPR16588 , 1:500), polyclonal goat antiIBA1 (Wako; 011-27991, 1:500), guinea pig antiIBA1 (Synaptic Systems; 234 308, clone Gp311H9, 1:1,000), rabbit polyclonal anti-ChAT (Sigma-Aldrich, AB143; 1:100), rabbit polyclonal anti-P2Y12 (AnaSpec; AS-55043A; 1:1,000), human anti-HuD ( ) (from patients with paraneoplastic neurological autoimmunity related to small-cell lung carcinoma; 1:10,000), polyclonal goat anti-human/mouse/rat Galectin-3 antibody (R&D Systems; AF1197, NP_034835 , 1: 1,000), and mouse anti-4-HNE (R&D Systems; MAB3249-SP, clone 198960, 1:200).

Techniques: Control, Flow Cytometry, Activation Assay

Journal: JCI Insight

Article Title: The CHI3L1-neutrophil axis drives immune suppression and breast cancer metastatic dissemination

doi: 10.1172/jci.insight.199307

Figure Lengend Snippet: ( A ) Staining of WT, Chi3l1 OE, Stat3 –/– , and Stat3 –/– Chi3l1 OE MIC mammary glands at 2 weeks after induction for CD8, granzyme B (GZMB), CD3, Pan-CK, and DAPI. ( B ) RNA FISH against IFN-γ with staining for CD3, Pan-CK, and DAPI on mammary tissue from WT, Chi3l1 OE, Stat3 –/– , and Stat3 –/– Chi3l1 OE MIC mammary glands at 2 weeks after induction. ( C – G ) Quantification of total CD3 + CD8 + , CD3 + CD8 + GZMB + , and tumor-infiltrating CD3 + CD8 + T cells, CD3 + IFN-γ + cells, and Ly6G + cells in WT ( n = 5), Chi3l1 OE ( n = 8), Stat3 –/– ( n = 6), and Stat3 –/– Chi3l1 OE ( n = 9) MIC mammary glands at 2 weeks after induction. ( H ) Staining of WT, Chi3l1 OE, Stat3 –/– , and Stat3 –/– Chi3l1 OE MIC mammary glands at 2 weeks after induction using antibodies against Ly6G, MPO, NE, pan-CK, and DAPI. ( I and J ) Quantification of total Ly6G + MPO + cells and Ly6G + MPO + NE + cells in WT ( n = 5), Chi3l1 OE ( n = 8), Stat3 –/– ( n = 6), and Stat3 –/– Chi3l1 OE ( n = 9) MIC mammary glands at 2 weeks after induction. ( K ) Quantification of CitH3 immunoblots normalized to α-tubulin. ( L ) Immunoblots for CitH3 and α-tubulin on WT ( n = 3), Chi3l1 OE ( n = 3), Stat3 –/– ( n = 3), and Stat3 –/– Chi3l1 OE ( n = 3) MIC mammary glands at 2 weeks after induction. * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001 by 1-way ANOVA with Tukey’s post hoc test. Scale bars: 100 μm.

Article Snippet: To deplete neutrophils, cohorts of MTB MIC CHI3L1 OE mice were randomized to receive 400 μg of purified anti-LY6G rat antibody (1A8, Bio X Cell BE0075-1) or 400 μg of IgG2a rat isotype control (2A3, Bio X Cell BE0089) via intraperitoneal (i.p.) injections twice weekly.

Techniques: Staining, Western Blot

Journal: JCI Insight

Article Title: The CHI3L1-neutrophil axis drives immune suppression and breast cancer metastatic dissemination

doi: 10.1172/jci.insight.199307

Figure Lengend Snippet: ( A ) H&E staining of mammary glands from IgG2a- and anti-Ly6G–treated MIC Stat3 –/– Chi3l1 OE mice at 2 weeks after induction. ( B ) Quantification of hyperplastic area in IgG2a-treated ( n = 8) and anti-Ly6G–treated ( n = 8) MIC Stat3 –/– Chi3l1 OE mice. Represented as percentage of mammary gland area. ( C ) Staining of mammary tissue from MIC Stat3 –/– Chi3l1 OE mice treated with anti-Ly6G or IgG2a for 2 weeks, for GZMB, CD8, CD3, pan-CK, and DAPI. ( D – F ) Quantification of CD3 + CD8 + , CD3 + CD8 + GZMB + , and tumor-infiltrating CD3 + CD8 + T cells in IgG2a-treated ( n = 8) and anti-Ly6G–treated ( n = 8) MIC Stat3 –/– Chi3l1 OE mammary glands. ( G ) Staining of mammary tissue from MIC Stat3 –/– Chi3l1 OE mice treated with anti-Ly6G or IgG2a, for PD-1, CD4, CD3, pan-CK, and DAPI. ( H – J ) Quantification of CD3 + CD4 + , CD3 + CD4 + PD-1 + , and tumor-infiltrating CD3 + CD4 + T cells in IgG2a-treated ( n = 8) and anti-Ly6G–treated ( n = 8) MIC Stat3 –/– Chi3l1 OE mammary glands. ( K ) H&E staining of mammary glands from IgG2a- and anti-Ly6G–treated MIC Chi3l1 OE mice at 2 weeks after induction. ( L ) Quantification of hyperplastic area in IgG2a-treated ( n = 6) and anti-Ly6G–treated ( n = 8) MIC Chi3l1 OE mice. Represented as percentage of mammary gland area. ( M ) Staining of mammary tissue from MIC Chi3l1 OE mice treated with anti-Ly6G or IgG2a for 2 weeks, for GZMB, CD8, CD3, pan-CK, and DAPI. ( N – P ) Quantification of CD3 + CD8 + , CD3 + CD8 + GZMB + , and tumor-infiltrating CD3 + CD8 + T cells in IgG2a-treated ( n = 6) and anti-Ly6G–treated ( n = 8) MIC Chi3l1 OE mammary glands. ( Q ) Staining of mammary tissue from MIC Chi3l1 OE mice treated with anti-Ly6G or IgG2a for 2 weeks, for PD-1, CD4, CD3, pan-CK, and DAPI. ( R – T ) Quantification of CD3 + CD4 + , CD3 + CD4 + PD1 + , and tumor-infiltrating CD3 + CD4 + T cells in IgG2a-treated ( n = 6) and anti-Ly6G–treated ( n = 8) MIC Chi3l1 OE mammary glands. * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001 by unpaired Student’s t test. Scale bars: 100 μm in A , C , G , M , and Q ; 500 μm in K .

Article Snippet: To deplete neutrophils, cohorts of MTB MIC CHI3L1 OE mice were randomized to receive 400 μg of purified anti-LY6G rat antibody (1A8, Bio X Cell BE0075-1) or 400 μg of IgG2a rat isotype control (2A3, Bio X Cell BE0089) via intraperitoneal (i.p.) injections twice weekly.

Techniques: Staining

Journal: JCI Insight

Article Title: The CHI3L1-neutrophil axis drives immune suppression and breast cancer metastatic dissemination

doi: 10.1172/jci.insight.199307

Figure Lengend Snippet: ( A ) H&E staining of WT, Chi3l1 OE, Stat3 –/– , and Stat3 –/– Chi3l1 OE MIC lungs at mammary tumor endpoint. ( B ) Percentage of WT ( n = 12), Chi3l1 OE ( n = 8), Stat3 –/– ( n = 13), and Stat3 –/– Chi3l1 OE ( n = 11) MIC mice with pulmonary metastases. ( C ) Quantification of metastatic area (percent total lung area) in WT ( n = 12), Chi3l1 OE ( n = 8), Stat3 –/– ( n = 13), and Stat3 –/– Chi3l1 OE ( n = 11) MIC lungs. ( D ) Quantification of PyMT + cells (percent live cells in blood) of WT ( n = 4) and Chi3l1 OE ( n = 3) MIC mice. ( E ) FACS for circulating PyMT + cells in WT and Chi3l1 OE MIC blood at 2 weeks after induction. ( F ) Staining of WT and Chi3l1 OE MIC mammary glands at 2 weeks after induction for laminin, collagen IV, pan-CK, and DAPI. ( G and H ) Quantification of area occupied by laminin or collagen IV (percent total mammary gland area) in WT ( n = 7) and Chi3l1 OE ( n = 8) MIC mammary glands. ( I ) FACS for circulating PyMT + cells in blood of Chi3l1 OE MIC mice after anti-Ly6G or IgG2a treatment for 2 weeks. ( J ) Quantification of PyMT + cells (percent live cells in the blood) of Chi3l1 OE MIC mice treated with IgG2a ( n = 6) or anti-Ly6G ( n = 5). ( K ) Staining of mammary tissue from MIC Chi3l1 OE mice treated with anti-Ly6G or IgG2a for laminin, collagen IV, pan-CK, and DAPI. ( L and M ) Quantification of area occupied by laminin and collagen IV (percent mammary gland area) in IgG2a-treated ( n = 6) and anti-Ly6G–treated ( n = 8) MIC Chi3l1 OE mammary glands. ( N ) Schematic of Transwell invasion assay through synthetic ECM. Created in BioRender (Muller W, 2026, https://BioRender.com/4lhz2a3 ). ( O ) ECM-invading PyMT + cells with PBS, rmChi3l1, or murine neutrophils, stained with crystal violet. Neutrophils + PBS (no PyMT cells) is shown as negative control. ( P ) Quantification of invading PyMT + cells. n = 3 technical replicates for each condition. * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001 by 1-way ANOVA with Tukey’s post hoc test ( C and P ) or by unpaired Student’s t test (for D , G , H , J , L , and M ). Scale bars: 1 cm in A , left panels; 1 mm in A , right panels; 100 μm in F , K , and O .

Article Snippet: To deplete neutrophils, cohorts of MTB MIC CHI3L1 OE mice were randomized to receive 400 μg of purified anti-LY6G rat antibody (1A8, Bio X Cell BE0075-1) or 400 μg of IgG2a rat isotype control (2A3, Bio X Cell BE0089) via intraperitoneal (i.p.) injections twice weekly.

Techniques: Staining, Transwell Invasion Assay, Negative Control

Journal: JCI Insight

Article Title: The CHI3L1-neutrophil axis drives immune suppression and breast cancer metastatic dissemination

doi: 10.1172/jci.insight.199307

Figure Lengend Snippet: ( A ) Immunoblots for Chi3l1, Stat3, and α-tubulin on WT ( n = 7) and Chi3l1 –/– ( n = 6) MIC mammary endpoint tumors. ( B ) Representative H&E staining of WT and Chi3l1 –/– MIC lungs at mammary tumor endpoint. ( C ) Percentage of WT ( n = 11) and Chi3l1 –/– ( n = 11) MIC mice with pulmonary metastases. ( D ) Quantification of total pulmonary metastatic area in WT ( n = 11) and Chi3l1 –/– ( n = 11) MIC lungs. Represented as percentage of total lung area. ( E ) Number of lung metastatic lesions in WT ( n = 11) and Chi3l1 –/– ( n = 11) MIC lungs. ( F ) Staining of WT and Chi3l1 –/– MIC mammary glands at 2 weeks after induction, for Ly6G, MPO, NE, pan-CK, and DAPI. ( G and H ) Quantification of Ly6G + cells and Ly6G + MPO + NE + cells in WT ( n = 7) and Chi3l1 –/– ( n = 10) MIC mammary glands at 2 weeks after induction. ( I ) Staining of WT and Chi3l1 –/– MIC mammary glands at 2 weeks after induction, for laminin, collagen IV, pan-CK, and DAPI. ( J and K ) Quantification of area occupied by laminin or collagen IV in WT ( n = 7) and Chi3l1 –/– ( n = 10) MIC mammary glands at 2 weeks after induction. Represented as percentage of total mammary gland area. ( L ) FACS sorting for PyMT + tumor cells in the blood of WT MIC mice treated with anti-Chi3l1 neutralizing antibody or IgG2b isotype control at 2 weeks after induction. ( M ) Quantification of PyMT + cells as percentage of total live cells in the blood of WT MIC mice treated with IgG2b ( n = 5) or anti-Chi3l1 ( n = 5) for 2 weeks. * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001 by unpaired Student’s t test. Scale bars: 1 mm in B , left panels; 100 μm in B , right panels; 10 μm in F and I .

Article Snippet: To deplete neutrophils, cohorts of MTB MIC CHI3L1 OE mice were randomized to receive 400 μg of purified anti-LY6G rat antibody (1A8, Bio X Cell BE0075-1) or 400 μg of IgG2a rat isotype control (2A3, Bio X Cell BE0089) via intraperitoneal (i.p.) injections twice weekly.

Techniques: Western Blot, Staining, Control

Journal: Science Advances

Article Title: Inflammation-triggered self-immolative conjugates enable oral peptide delivery by overcoming gastrointestinal barriers

doi: 10.1126/sciadv.aea2989

Figure Lengend Snippet: ( A ) Representative trajectories of Cy5-proKPV NPs and Cy5-labeled PS NPs in the mouse intestinal mucus, as determined by MTrackJ analysis of 5-min confocal microscopy recordings. ( B ) Time-dependent ensemble-averaged geometric MSD of proKPV NPs and PS NPs in the mouse intestinal mucus. ( C ) Distributions of log( D eff ) for proKPV NPs and PS NPs at a 5-min timescale, with at least 100 particles tracked per sample for each type. ( D ) Three-dimensional fluorescence images illustrate the penetration of Cy5-KPV and Cy5-proKPV NPs through the mouse intestinal mucus. ( E ) Representative ex vivo images (left) and quantitative analysis (right) illustrating accumulation of Cy5-labeled proKPV in the colons of mice with or without DSS-induced acute colitis at 6 hours after oral administration. ( F and G ) Fluorescence images (F) and quantitative analysis (G) of the colonic tissues isolated from colitis mice at different time points following treatment with free Cy5-KPV or Cy5-proKPV. ( H ) AUC of Cy5 fluorescence in colonic tissues after treatment with free Cy5-KPV or Cy5-proKPV. ( I ) Fluorescence images of cryosections of colonic tissues from colitis mice at 6 hours after treatment with Cy5-KPV or Cy5-proKPV. Nuclei were stained with DAPI (blue). ( J to L ) Immunofluorescence analysis of colocalization of Cy5-KPV or Cy5-proKPV with CK18 + intestinal epithelial cells (J), CD68 + macrophages (K), or Ly6G + neutrophils (L) in cryosections of colonic tissues. Data in (E), (G), and (H) are presented as means ± SD ( n = 3 biological replicates). * P < 0.05; *** P < 0.001.

Article Snippet: The obtained cryosections were separately stained with anti–cytokeratin 18 (CK18) antibody (bsm-52058R, Bioss Biotechnology, China), anti-F4/80 antibody (30325, Cell Signaling Technology, USA), or anti-Ly6G antibody (31469, Cell Signaling Technology, USA) to further evaluate the tissue distribution of NPs.

Techniques: Labeling, Confocal Microscopy, Fluorescence, Ex Vivo, Isolation, Staining, Immunofluorescence

Journal: Science Advances

Article Title: Inflammation-triggered self-immolative conjugates enable oral peptide delivery by overcoming gastrointestinal barriers

doi: 10.1126/sciadv.aea2989

Figure Lengend Snippet: ( A ) Schematic illustration of treatment regimens. ( B ) Body weight of mice during a 7-day treatment course. Data were normalized as the percentages of the body weight at day 0.** P < 0.01; *** P < 0.001; n.s., not significant versus the colitis group. ( C ) Changes in DAI. ** P < 0.01 and *** P < 0.001 versus the colitis group. ( D ) Digital photos (left) and quantified lengths (right) of colonic tissues isolated from mice at day 7 following different treatments. Scale bars, 5 mm. ( E ) Representative miniendoscopic images of colons from mice after 7 days of treatment. ( F ) Colonic tissue sections stained with H&E or PAS for different groups. ( G ) Immunofluorescence analysis of the expression of CK18, occludin, and ZO-1 in colonic tissues from mice following various treatments. Scale bars, 100 μm. ( H to M ) Levels of TNF-α (H), IL-1β (I), IL-6 (J), MPO (K), MDA (L), and ROS (M) in colonic tissues isolated from healthy or diseased mice treated with different formulations. After 7 days of treatment, colonic tissue homogenates were prepared, and mediator concentrations were quantified and normalized to the total protein content. ( N ) Immunofluorescence of colonic tissues showing Ly6G + neutrophils, CD206 + M2 macrophages, CitH3/NE-positive NETs, and FOXP3 + T reg cells after different treatments. Data in (B) to (D) and (H) to (M) are presented as means ± SD ( n = 6 biological replicates). * P < 0.05; ** P < 0.01; *** P < 0.001; n.s., not significant.

Article Snippet: The obtained cryosections were separately stained with anti–cytokeratin 18 (CK18) antibody (bsm-52058R, Bioss Biotechnology, China), anti-F4/80 antibody (30325, Cell Signaling Technology, USA), or anti-Ly6G antibody (31469, Cell Signaling Technology, USA) to further evaluate the tissue distribution of NPs.

Techniques: Isolation, Staining, Immunofluorescence, Expressing

Journal: Science Advances

Article Title: Inflammation-triggered self-immolative conjugates enable oral peptide delivery by overcoming gastrointestinal barriers

doi: 10.1126/sciadv.aea2989

Figure Lengend Snippet: ( A ) Body weight changes of mice during 7-day treatment, which were normalized to the baseline at day 0. ** P < 0.01 and *** P < 0.001; n.s., not significant versus the colitis group. ( B ) Changes in DAI values. ** P < 0.01 and *** P < 0.001 versus the colitis group. ( C and D ) Digital photos (C) and quantified lengths (D) of colonic tissues isolated from mice after 7 days of treatment. ( E ) Histological sections of colonic tissues stained with H&E or PAS. ( F ) Immunofluorescence indicates expression patterns of CK18, occludin, and ZO-1 in colonic tissues. ( G to J ) Levels of TNF-α (G), IL-6 (H), IL-1β (I), and MPO (J) in colonic tissues isolated from healthy or diseased mice treated with different formulations. ( K and L ) Immunofluorescence analysis of NETs (K), Ly6G + neutrophils, and CD206 + macrophages (L) in colonic tissues. Data in (A), (B), (D), and (G) to (J) are presented as means ± SD ( n = 6 biological replicates). * P < 0.05; ** P < 0.01; *** P < 0.001; n.s., not significant.

Article Snippet: The obtained cryosections were separately stained with anti–cytokeratin 18 (CK18) antibody (bsm-52058R, Bioss Biotechnology, China), anti-F4/80 antibody (30325, Cell Signaling Technology, USA), or anti-Ly6G antibody (31469, Cell Signaling Technology, USA) to further evaluate the tissue distribution of NPs.

Techniques: Isolation, Staining, Immunofluorescence, Expressing

Journal: bioRxiv

Article Title: Neutrophil terminal programming in the ischemic heart drives fibrosis after myocardial infarction

doi: 10.64898/2026.01.15.699253

Figure Lengend Snippet: Targeting Ly6G modulates the immune cell landscape in the heart after MI with increased pro-fibrotic γδ T cells. a) UMAP visualization of total heart CD45 + cells 5 days post MI, in mice with or without Ly6G targeting (see ). b) Cell count per mg of heart tissue for the indicated cell type clusters identified in (a). c) Macrophage content in the infarct area 14 days post-MI in isotype or anti-Ly6G treated groups. Left panel: immunofluorescence staining of CD68 (pink) on heart sections, scale bar = 200µm. Right panel: quantification of macrophage area expressed as percentage of the infarcted area. d) CellChat analysis of γδ T cells communication with cardiac fibroblasts, probability of communication pathways (ligand-receptor pairs) e) Scratch assay of 3T3 fibroblasts incubated 24h with activated γδ T cells, representative images at t=0, t=18h and t=24h (left panel) and quantification of percentage of wound closure after 18h and 24h (right panel). Each point represents one well, pooled from three independent experiments. f) Proposed model and summary of the main findings. Bar plots: one circle represents one mouse, open circles represent female mice. MI: myocardial infarction, CTL: control, Macro: Macrophages, Prolif: Proliferating, Neutro: Neutrophils, DC: Dendritic Cells, cDC1: conventional Dendritic Cell 1, DC: Dendritic Cells, pDC: plasmacytoid Dendritic Cells, NI: non-identified. Statistical tests: (panel b,c,e) unpaired t-test (*<0.05; **<0.01). Illustration (panel f) created with Biorender.

Article Snippet: For Ly6G targeting, mice received daily intraperitoneal injections of 25μg of rat IgG2aκ anti-mouse Ly6G InVivoPlusTM (BioXCell, BP0075-1) + 50μg of mouse anti-rat IgG2aκ (BioXCell, BE0122) (anti-Ly6G group), and the control group received 25μg of rat IgG2aκ isotype control (BioXCell, BP0089) + 50μg of mouse anti-rat IgG2aκ (BioXCell, BE0122) (isotype group).

Techniques: Cell Characterization, Immunofluorescence, Staining, Wound Healing Assay, Incubation, Control

Journal: bioRxiv

Article Title: Neutrophil terminal programming in the ischemic heart drives fibrosis after myocardial infarction

doi: 10.64898/2026.01.15.699253

Figure Lengend Snippet: a) UMAP plot visualization of heart neutrophils extracted from and reclustered. b) UMAP plot from panel a) split by timepoint after MI. c) Percentage of heart neutrophil clusters in total heart neutrophils according to time post MI. One circle represents one mouse, open circles represent female mice. d) Relative expression of Csf2ra , Tgfbr1 and Tgfbr2 transcripts in total neutrophils in the indicated conditions. e) relative gene expression in neutrophils left untreated or GM-CSF+TGFβ stimulated neutrophils assessed by quantitative PCR (n= neutrophils from 2 mice). f) CITE-seq signal of the indicated surface protein on bone marrow, spleen, blood and heart neutrophils at day 4 post-MI projected onto the UMAP plot from . g) Surface protein expression of the indicated markers on pre-gated live Ly6G+ neutrophils assessed by flow cytometry, in unstimulated bone marrow-isolated neutrophil, and GM-CSF+TGFβ stimulated neutrophils. MI: myocardial infarction, BM: bone marrow, CTL: control. Statistical tests: (panel c) unpaired t-test (*<0.05; **<0.01; ***<0.001; ****<0.0001).

Article Snippet: For Ly6G targeting, mice received daily intraperitoneal injections of 25μg of rat IgG2aκ anti-mouse Ly6G InVivoPlusTM (BioXCell, BP0075-1) + 50μg of mouse anti-rat IgG2aκ (BioXCell, BE0122) (anti-Ly6G group), and the control group received 25μg of rat IgG2aκ isotype control (BioXCell, BP0089) + 50μg of mouse anti-rat IgG2aκ (BioXCell, BE0122) (isotype group).

Techniques: Expressing, Gene Expression, Real-time Polymerase Chain Reaction, Flow Cytometry, Isolation, Control

Journal: bioRxiv

Article Title: Neutrophil terminal programming in the ischemic heart drives fibrosis after myocardial infarction

doi: 10.64898/2026.01.15.699253

Figure Lengend Snippet: a) Experimental design of Ly6G targeting experiments. b) Flow cytometry analysis of heart neutrophils 3 days post MI: percentage of Neutrophils among total CD45 + cells (left panel) and percentage of SiglecF + neutrophils among total neutrophils (right panel). c) Interstitial fibrosis in the infarct border zone 14 days post-MI, expressed as percentage of total area, determined by picrosirius red staining, scale bar: 100µm. d) Experimental design of the single cell RNA-seq/CITE-seq experiments with Ly6G targeting. Group isotype MI n= 4 females and 6 males, group anti-Ly6G MI n= 5 females and 10 males, group isotype sham n= 3 males and 3 females, group anti-Ly6G sham n= 3 males and 3 females. e) UMAP plot of neutrophils identified in (Supplementary Figure 4a), extracted from the MI groups and reclustered. N=3 independent scRNA-seq/CITE-seq experiments were integrated. f) Relative expression of the main marker genes used to identify neutrophil subsets. g) Neutrophil counts per mg of heart tissue at day 5 post-MI. h) UMAP plot of neutrophils from panel e) split by treatment. i) Neutrophil cluster proportions expressed as percentage of total neutrophils. j) Gene count for 5 of the top16 up regulated biological processes in the Mature Retnlg hi cluster vs. the Mature cluster (green bars) and in the SiglecF + Ly6G lo cluster vs. the all SiglecF+ neutrophil clusters (pink bars); all with adjusted p-value <0.05 k) Expression of the indicated scores by neutrophil clusters from panel e). Bar plots: one circle represents one mouse, open circles represent female mice. MI: myocardial infarction, BM: bone marrow, Iso: isotype, GO: gene ontology. Statistical tests: (panel b, c, g, i) unpaired t-test (*<0.05; **<0.01; ***<0.001; ****<0.0001). Illustration (panel a,d) created with Biorender.

Article Snippet: For Ly6G targeting, mice received daily intraperitoneal injections of 25μg of rat IgG2aκ anti-mouse Ly6G InVivoPlusTM (BioXCell, BP0075-1) + 50μg of mouse anti-rat IgG2aκ (BioXCell, BE0122) (anti-Ly6G group), and the control group received 25μg of rat IgG2aκ isotype control (BioXCell, BP0089) + 50μg of mouse anti-rat IgG2aκ (BioXCell, BE0122) (isotype group).

Techniques: Flow Cytometry, Staining, RNA Sequencing, Expressing, Marker

Journal: bioRxiv

Article Title: Neutrophil terminal programming in the ischemic heart drives fibrosis after myocardial infarction

doi: 10.64898/2026.01.15.699253

Figure Lengend Snippet: a) Expression of the indicated scores in control and day 5, 7 and 14 post-MI heart cells from an in-house scRNA-seq atlas. b) Immunofluorescent staining of day 3 post-MI heart section showing the infarct and border zone area. Hoechst is stained in blue, Ly6G (neutrophils) in green and PDGFRα (fibroblasts) in red, scale bar: 50µm. c) Immunofluorescence staining of PDGFRα in the peri infarct region 3 days post-MI, expressed as PDGFRα + area percentage of total area (left panel). PDGFRα is stained in red and Hoechst in blue (right panel), scale bar: 100µm. d) Scratch assay on 3T3 fibroblasts incubated 24h with untreated or SiglecF + neutrophils; representative images at t=0 and t=24h (left panel), quantification of percentage of wound closure after 24h (right panel); each point represents one well, pooled from three independent experiments. e,f) CellChat analysis of heart neutrophil cluster (from ) communication to cardiac fibroblasts; e) Probability of communication pathways (ligand-receptor pairs); f) associated CypA and TGFβ signaling pathway network. g,h) Percentage of wound closure after 18h and 24h from scratch assay of 3T3 fibroblasts incubated 24h with SiglecF + neutrophils and Galunisertib g) or CypA inhibitor h). Each point represents one well, pooled from two independent experiments. MI: myocardial infarction, Fibro: Fibroblasts, Myo Fibro: Myofibroblatsts, VSMC: Vascular smooth muscle cell, EC: Endothelial cell, Macro: Macrophage, cDC: conventional dendritic cell, pDC: plasmacytoid dendritic cell, Neutro: Neutrophil, Mast: Mastocyte, Baso: Basophil, T: T cell, gdT: gamma delta T cell, NK: Natural Killer, ILC: innate lymphoid cell, ECM: extracellular matrix, Glycoprot: glycoproteins, prot: proteins. Statistical tests: (panel c) unpaired t-test, (panel d,g,h) Ordinary one-way ANOVA (Fisher’s LSD test) (*<0.05; **<0.01; ***<0.001).

Article Snippet: For Ly6G targeting, mice received daily intraperitoneal injections of 25μg of rat IgG2aκ anti-mouse Ly6G InVivoPlusTM (BioXCell, BP0075-1) + 50μg of mouse anti-rat IgG2aκ (BioXCell, BE0122) (anti-Ly6G group), and the control group received 25μg of rat IgG2aκ isotype control (BioXCell, BP0089) + 50μg of mouse anti-rat IgG2aκ (BioXCell, BE0122) (isotype group).

Techniques: Expressing, Control, Staining, Immunofluorescence, Wound Healing Assay, Incubation

Journal: Cellular and Molecular Gastroenterology and Hepatology

Article Title: ATG16L1 Regulates Reparative Function of Peritoneal Macrophages During Acute Drug-induced Liver Injury

doi: 10.1016/j.jcmgh.2025.101674

Figure Lengend Snippet: Intrahepatic migration of PMs promotes liver repair post-DILI. Liver tissues from APAP-treated mice were collected at 24 hours post DILI. F4/80 and GATA6 co-staining was analyzed by IHC ( A ) and IF ( B ). PMs were isolated and intraperitoneally injected into APAP-pretreated mice or depleted prior to APAP-induced DILI. Liver tissues and serum were collected at 24, 48, and 72 hours post DILI. Representative H&E ( C ), TUNEL ( D ), CD31 ( E ) staining, serum ALT ( F ) and AST ( G ), IL-1β ( H–I ), and IL-10 ( J–K ) levels were analyzed; furthermore, representative IF staining of CD11b ( L ) and Ly6G ( M ) was analyzed. n = 6/group. Data are shown as mean ± SEM. Each point represents an independent experiment. ∗ P < .05, ∗∗ P < .01, ∗∗∗ P < .001, ∗∗∗∗ P < .0001.

Article Snippet: For immunohistochemical (IHC) and immunofluorescence (IF) staining, the sections were dehydrated and antigens were retrieved by incubation overnight at 4°C with the following primary antibodies: rat anti-mouse F4/80 (1:100, ab11101, Abcam), mouse anti-mouse GATA6 (1:200, 67943-1-Ig, proteintech), rabbit anti-mouse ATG16L1 (1:200, ab188642, Abcam), rabbit anti-mouse platelet endothelial cell adhesion molecule-1 (CD31) (1:100, ab222783, Abcam), hyaluronan-binding protein 1/complement component 1, q subcomponent binding protein (HABP1/C1QBP) (HY- P76185 , MCE), Mouse (His) (1:100, ab213204, Abcam), rabbit anti-mouse LC3 (1:200, ab192890, Abcam), rat anti-mouse CD11b (1:50, ab8878, Abcam), rat anti-mouse Ly6G (1:500, 88876, CST), rat anti-mouse Clec4f (MAB2784; 1/1000, R&D Systems), and rabbit anti-mouse PCNA (1:200, ab29, Abcam) antibodies.

Techniques: Migration, Staining, Isolation, Injection, TUNEL Assay

Journal: Cellular and Molecular Gastroenterology and Hepatology

Article Title: ATG16L1 Regulates Reparative Function of Peritoneal Macrophages During Acute Drug-induced Liver Injury

doi: 10.1016/j.jcmgh.2025.101674

Figure Lengend Snippet: ATG16L1 influences the pro-repair effects of PMs. Liver tissues from APAP-treated mice were collected at 24 hours post DILI. Immunostaining for F4/80, GATA6, ATG16L1, and DAPI ( A ). CD45.2 PMs were injected intraperitoneally into CD45.1 mice that had been pretreated with APAP. After 24 hours, the intrahepatic NPCs were collected. To isolate the migrated CD45.2 macrophages specifically, they were purified from the liver sample using CD45.2-based magnetic sorting. Stable CD45.2 PMs were used as the control group. The samples were analyzed by flow cytometry ( B ) and WB for ATG16L1 ( C ); n = 3/group. WB analysis of ATG16L1 in peritoneal macrophages ( D ). PMs were injected into APAP-pretreated WT mice, and liver tissues/serum were collected at 0, 24, 48, and 72 hours post DILI. H&E ( E ), TUNEL ( F ), CD31 ( G ), ALT ( H ), AST ( I ), IL-1β ( J–K ), and IL-10 ( L–M ) analyses. Furthermore, representative IF staining of CD11b ( N ) and Ly6G ( O ) was evaluated. n = 6/group. Data are shown as mean ± SEM. Each point represents an independent experiment. ∗ P < .05, ∗∗ P < .01, ∗∗∗∗ P < .0001.

Article Snippet: For immunohistochemical (IHC) and immunofluorescence (IF) staining, the sections were dehydrated and antigens were retrieved by incubation overnight at 4°C with the following primary antibodies: rat anti-mouse F4/80 (1:100, ab11101, Abcam), mouse anti-mouse GATA6 (1:200, 67943-1-Ig, proteintech), rabbit anti-mouse ATG16L1 (1:200, ab188642, Abcam), rabbit anti-mouse platelet endothelial cell adhesion molecule-1 (CD31) (1:100, ab222783, Abcam), hyaluronan-binding protein 1/complement component 1, q subcomponent binding protein (HABP1/C1QBP) (HY- P76185 , MCE), Mouse (His) (1:100, ab213204, Abcam), rabbit anti-mouse LC3 (1:200, ab192890, Abcam), rat anti-mouse CD11b (1:50, ab8878, Abcam), rat anti-mouse Ly6G (1:500, 88876, CST), rat anti-mouse Clec4f (MAB2784; 1/1000, R&D Systems), and rabbit anti-mouse PCNA (1:200, ab29, Abcam) antibodies.

Techniques: Immunostaining, Injection, Purification, Control, Flow Cytometry, TUNEL Assay, Staining