Journal: Frontiers in Cardiovascular Medicine

Article Title: Deficiency of Myeloid Pfkfb3 Protects Mice From Lung Edema and Cardiac Dysfunction in LPS-Induced Endotoxemia

doi: 10.3389/fcvm.2021.745810

Figure Lengend Snippet: Myeloid-specific Pfkfb3 deficiency attenuates LPS-induced inflammatory responses. (A) Representative images (left) and quantification (right) for immunohistochemical staining of the neutrophil marker Ly6G in lung sections of Pfkfb3 WT mice and Pfkfb3 ΔMϕ mice 6 h after LPS injection ( n = 5). (B) Representative images (left) and quantification (right) for immunohistochemical staining of the macrophage marker Mac2 in lung sections of Pfkfb3 WT mice and Pfkfb3 ΔMϕ mice 6 h after LPS injection ( n = 5). (C–E) qPCR analysis of the mRNA levels of Il1b (C) , Il6 (D) and Nos2 (E) in the lung of Pfkfb3 WT mice and Pfkfb3 ΔMϕ mice 6 h after LPS injection ( n = 6). (F,G) ELISA analysis of Il1b (F) and Il6 (G) in serum of Pfkfb3 WT mice and Pfkfb3 ΔMϕ mice 6 h after LPS injection ( n = 4). (H) NO levels in serum of Pfkfb3 WT mice and Pfkfb3 ΔMϕ mice 6 h after LPS injection ( n = 3–4). All data are represented as mean ± SEM, ** P < 0.01 and *** P < 0.001 for Pfkfb3 WT vs. Pfkfb3 ΔMϕ (unpaired two-tailed Student's t test).

Article Snippet: After antigen retrieval with Antigen Unmasking Solution (H-3301, Vector Laboratories, Burlingame, CA, USA) at 98°C for 10 min, sections were blocked with avidin solution with 10% normal rabbit serum for 1 h at room temperature, and incubated in biotin blocking solution with primary antibodies against Mac2 (3 μg/mL, CL8942F, Cedarlane, Burlington, NC, USA), or Ly6G (3 μg/mL, 551459, BD biosciences, San Jose, CA, USA) at 4°C overnight.

Techniques: Immunohistochemical staining, Staining, Marker, Injection, Enzyme-linked Immunosorbent Assay, Two Tailed Test

Journal: Communications Biology

Article Title: TIM-3 on myeloid cells promotes pulmonary inflammation through increased production of galectin-3

doi: 10.1038/s42003-024-06762-w

Figure Lengend Snippet: a Bulk RNA-sequencing data obtained from whole lungs of FSF-TIM3 (WT) and FSF-TIM3/ LysM -Cre +/− mice, visualized with a Volcano plot highlighting differentially expressed genes (DEGs) ( n = 3 per group, log 2 FC ≥ 2, P < 0.05). b Heatmap showing the expression of galectins and galectin-binding protein. Data shown are relative to the Z-scores calculated across the samples. Red indicates relatively high levels of expression; blue indicates relatively low levels of expression. c Protein extracts were obtained from the lung tissues of WT and FSF-TIM3/ LysM -Cre +/+ mice at 7 and 14 weeks of age. The samples were subjected to western blot analysis with the indicated antibodies. d Western blot analysis was carried out on BALF supernatants with an anti-galectin-3 antibody. e ELISA was performed to assess the concentration of galectin-3 in BALF ( n = 5 per group). Graphs show the mean ± SD and the data were analyzed by two-way ANOVA with Sidak’s multiple comparison test. **P < 0.01 and ****P < 0.0001 .

Article Snippet: For galectin-3 inhibition, 1 mg/kg galectin-3 inhibitor GB1107 (1978336-61-6, MedChemExpress) per mouse or an equivalent dose of the vehicle was administered by intraperitoneal injection in 100 μl PBS every other day.

Techniques: RNA Sequencing, Expressing, Binding Assay, Western Blot, Enzyme-linked Immunosorbent Assay, Concentration Assay, Comparison

Journal: Communications Biology

Article Title: TIM-3 on myeloid cells promotes pulmonary inflammation through increased production of galectin-3

doi: 10.1038/s42003-024-06762-w

Figure Lengend Snippet: a Representative immunohistochemical images of lung tissues obtained from WT and FSF-TIM3/ LysM -Cre +/+ mice using the indicated antibodies ( n = 3 per group). b The relative and mean intensities were quantified using the Zen software. Graphs show the mean ± SD and the data were analyzed by one-way ANOVA with Tukey’s multiple comparison test. **P < 0.01 . c BALF was isolated from 4-week-old WT and FSF-TIM3/ LysM -Cre +/− mice ( n = 3 per group). Immunocytochemical analysis of BALF cells was performed using antibodies against TIM-3 (green) and galectin-3 (red), with nuclei counterstaining (DAPI). Quantification of mean fluorescence intensity was performed using the Zen software. d ELISA was applied to detect the concentration of galectin-3 in serum-free media of cultured BALF cells obtained from 4-week-old mice ( n = 4 per group). e Flow cytom e tric analysis of TIM-3 and galectin-3 in AMФ cells from BALF of 4-week-old mice. f Representative dot plots and histogram of TIM-3 and galectin-3 in AMФ cells from lung tissues obtained from WT and FSF-TIM3/ LysM -Cre +/− mice of the indicated age ( n = 4 per group). g Relative transcript levels of representative inflammation-associated cytokines were determined by RT-PCR in AMФ cells sorted from the lungs of 4-week-old WT and FSF-TIM3/ LysM -Cre +/− mice. AMФ cells were sorted from four mice and pooled into one sample (P). Graphs shows the mean ± SD and the data were analyzed by unpaired two-tailed Student’s t test. *P < 0.05 and ****P < 0.0001 .

Article Snippet: For galectin-3 inhibition, 1 mg/kg galectin-3 inhibitor GB1107 (1978336-61-6, MedChemExpress) per mouse or an equivalent dose of the vehicle was administered by intraperitoneal injection in 100 μl PBS every other day.

Techniques: Immunohistochemical staining, Software, Comparison, Isolation, Fluorescence, Enzyme-linked Immunosorbent Assay, Concentration Assay, Cell Culture, Reverse Transcription Polymerase Chain Reaction, Two Tailed Test

Journal: Communications Biology

Article Title: TIM-3 on myeloid cells promotes pulmonary inflammation through increased production of galectin-3

doi: 10.1038/s42003-024-06762-w

Figure Lengend Snippet: a Scheme of experiments used to generate the data shown in ( b – e ). FSF-TIM3/ LysM -Cre +/− mice were intraperitoneally treated with TIM-3-blocking antibody or control IgG (100 μg in 100 μl). ( n = 5 for 8-week-old mice, n = 7 for 12-week-old mice). b Representative images of the lungs treated with TIM-3-blocking antibody or IgG. Lung weights are shown in the graph. c Representative H&E staining images of lung tissues from IgG- or TIM-3 antibody-treated FSF-TIM3/ LysM -Cre +/− mice ( n = 3 per group). Graphs show the mean ± SD and the data were analyzed by two-way ANOVA with Sidak’s multiple comparison test. **P < 0.01 and ***P < 0.001 . d Western blot analysis was performed on lung tissues from IgG- or TIM-3 antibody-treated FSF-TIM3/ LysM -Cre +/− mice using the antibodies against galectin-3 and β-Actin. Graphs show the mean ± SD and the data were analyzed by unpaired two-tailed Student’s t-test. *P < 0.05 , and **P < 0.01 . e Cells were isolated from the lungs of 12-w e ek-old FSF-TIM3/ LysM -Cre +/− mice treated with a TIM-3-blocking antibody or control IgG, and then subjected to flow cytometry using antibodies against galectin-3 and representative marker antibodies for each cell type in the lungs.

Article Snippet: For galectin-3 inhibition, 1 mg/kg galectin-3 inhibitor GB1107 (1978336-61-6, MedChemExpress) per mouse or an equivalent dose of the vehicle was administered by intraperitoneal injection in 100 μl PBS every other day.

Techniques: Blocking Assay, Control, Staining, Comparison, Western Blot, Two Tailed Test, Isolation, Flow Cytometry, Marker

Journal: Communications Biology

Article Title: TIM-3 on myeloid cells promotes pulmonary inflammation through increased production of galectin-3

doi: 10.1038/s42003-024-06762-w

Figure Lengend Snippet: a Scheme of experiments used to generate the data shown in ( b – h ). FSF-TIM3/ LysM -Cre +/− mice were intraperitoneally treated with either GB1107 (1 mg/kg/every 2 days) or a vehicle control. b Representative images and weights of lungs from FSF-TIM3/ LysM -Cre +/− mice treated with GB1107 or the vehicle ( n = 6 for vehicle group, n = 7 for GB1107 group). c Representative H&E staining images of lung tissues from both vehicle- and GB1107-treated FSF-TIM3/ LysM -Cre +/− mice. d – g Quantitative analysis of total cell numbers ( d ), tissue area ( e ), cell density ( f ), and airspace area ( g ) in H&E-stained lung sections. Quantification was performed using InForm2.2.1 software, with at least three sections analyzed per slide. h Relative transcript levels of IL-1β and TNF-α determined by RT-PCR in FSF-TIM3/ LysM -Cre +/− mice ( n = 3 per group). Data are presented as mean ± SD and were analyzed using an unpaired two-tailed Student’s t-test. *P < 0.05 and **P < 0.01 .

Article Snippet: For galectin-3 inhibition, 1 mg/kg galectin-3 inhibitor GB1107 (1978336-61-6, MedChemExpress) per mouse or an equivalent dose of the vehicle was administered by intraperitoneal injection in 100 μl PBS every other day.

Techniques: Control, Staining, Software, Reverse Transcription Polymerase Chain Reaction, Two Tailed Test

Journal: Communications Biology

Article Title: TIM-3 on myeloid cells promotes pulmonary inflammation through increased production of galectin-3

doi: 10.1038/s42003-024-06762-w

Figure Lengend Snippet: a Conditional myeloid-specific TIM-3 knock-out mice (TIM3 f/f / LysM -Cre +/+ , TIM3-cKO) were generated by crossing the TIM3 f/f mice with LysM -Cre mice. b Schematic representation of the LPS-induced murine lung injury model used in this study; TIM3 f/f and TIM3 f/f / LysM -Cre +/+ mice were intranasally (i.n) injected with LPS (10 μg in 30 μl) or PBS and assessed after the indicated durations ( n = 3 for PBS group, n = 5 for LPS group). c Representative H&E staining images of lung tissues from the LPS-treated mice. d Inflammation scores based on histological examination of lung sections ( n = 5 per group). e Relative mRNA levels of TNFA and IL-1β in lung tissues of TIM3 f/f and TIM3 f/f / LysM -Cre +/+ mice at 72 h post LPS treatment were determined by qRT-PCR ( n = 3 per group). f Representative histogram of galectin-3 expression in myeloid cells from lung tissues of TIM3 f/f and TIM3 f/f / LysM -Cre +/+ mice at 72 h after intranasally administration of LPS ( n = 3 per group). Graphs show the mean ± SD and the data were analyzed by two-way ANOVA with Sidak’s multiple comparison test. *P < 0.05 , **P < 0.01 .

Article Snippet: For galectin-3 inhibition, 1 mg/kg galectin-3 inhibitor GB1107 (1978336-61-6, MedChemExpress) per mouse or an equivalent dose of the vehicle was administered by intraperitoneal injection in 100 μl PBS every other day.

Techniques: Knock-Out, Generated, Injection, Staining, Quantitative RT-PCR, Expressing, Comparison

Journal: The Journal of Clinical Investigation

Article Title: Itaconate suppresses atherosclerosis by activating a Nrf2-dependent antiinflammatory response in macrophages in mice

doi: 10.1172/JCI173034

Figure Lengend Snippet: WT and Acod1 –/– mice were induced to become atherosclerotic via PCSK9-AAV administration followed by 10-week Western diet. ( A and B ) Representative images of H&E-stained ( A ) aortic root and ( B ) brachiocephalic artery (BCA) sections of WT and Acod1 –/– mice. Arrows indicate atherosclerotic lesions and arrowheads indicate necrotic cores. ( C ) The quantifications of lesion area and necrotic area in each section of aortic root ( n = 23–24/group) and BCA ( n = 10/group) are shown. ( D – F ) The quantification of Mac2-positive area in each section of aortic root ( n = 13/group) and BCA ( n = 10/group) of atherosclerotic WT and Acod1 –/– mice is shown ( D ), with representative images of anti-Mac2–stained ( E ) aortic root and ( F ) BCA sections. ( G and H ) The inflammatory cytokines and chemokines’ ( G ) gene expression in atherosclerotic aorta and ( H ) protein levels in tissue culture medium of atherosclerotic aortas from WT and Acod1 –/– mice, including IL-1β, IL-6, IL-12, CCL2, CCL3, CCL5, CXCL1, CXCL2, and CXCL10, were measured by qRT-PCR ( n = 7/group) and multiplex assay ( n = 8–9/group), respectively. Results are presented as mean ± SEM. Two-way ANOVA followed by Tukey’s post hoc test was used in C and D and unpaired, 2-tailed Student’s t test was used in G and H for statistical analysis. Scale bars: 200 μm ( A and E ) and 50 μm ( B and F ).

Article Snippet: Immunohistochemical staining was performed to detect the macrophage marker Mac2 (sc-81728, Santa Cruz Biotechnology), and nuclei were counterstained with hematoxylin.

Techniques: Western Blot, Staining, Gene Expression, Quantitative RT-PCR, Multiplex Assay

Journal: The Journal of Clinical Investigation

Article Title: Itaconate suppresses atherosclerosis by activating a Nrf2-dependent antiinflammatory response in macrophages in mice

doi: 10.1172/JCI173034

Figure Lengend Snippet: Acod1 fl/fl and Acod1 fl/fl LysM cre mice were induced to become atherosclerotic via PCSK9-AAV administration followed by 10-week Western diet. ( A and B ) Representative images of H&E-stained ( A ) aortic root and ( B ) brachiocephalic artery (BCA) sections of atherosclerotic Acod1 fl/fl and Acod1 fl/fl LysM cre mice. Arrows indicate atherosclerotic lesions and arrowheads indicate necrotic cores. ( C ) The quantifications of lesion area and necrotic area in each section of aortic root ( n = 17–20/group) and BCA ( n = 17–19/group) are shown. ( D – F ) The quantification of Mac2-positive area in each section of aortic root ( n = 9/group) and BCA ( n = 11–13/group) of atherosclerotic Acod1 fl/fl and Acod1 fl/fl LysM cre mice is shown ( D ), along with representative images of anti-Mac2–stained ( E ) aortic root and ( F ) BCA sections. Results are shown as mean ± SEM. Two-way ANOVA followed by Tukey’s post hoc test was used for statistical analysis. P values indicate the main effect of the comparison. Scale bars: 200 μm ( A and E ) and 50 μm ( B and F ).

Article Snippet: Immunohistochemical staining was performed to detect the macrophage marker Mac2 (sc-81728, Santa Cruz Biotechnology), and nuclei were counterstained with hematoxylin.

Techniques: Western Blot, Staining, Comparison

Journal: The Journal of Clinical Investigation

Article Title: Itaconate suppresses atherosclerosis by activating a Nrf2-dependent antiinflammatory response in macrophages in mice

doi: 10.1172/JCI173034

Figure Lengend Snippet: ( A ) Representative images of H&E-stained aortic root and brachiocephalic artery (BCA) sections of mice with indicated treatment. Vehicle, vehicle control; OI, 4-octyl itaconate; Athero+Vehicle, atherosclerosis and vehicle; Athero+OI, atherosclerosis and 4-octyl itaconate. Arrows indicate atherosclerotic lesions and arrowheads indicate necrotic cores. ( B and C ) The quantifications of lesion area and necrotic area ( B ) as well as Mac2-positive area ( C ) in each section of aortic root and BCA from indicated mice are shown ( n = 7–8/group). ( D ) Representative images of anti-Mac2–stained aortic root and BCA sections of mice with indicated treatment. Results are shown as mean ± SEM. Two-way ANOVA followed by Tukey’s post hoc test was used for statistical analysis. P values indicate the main effect of the comparison between Athero+Vehicle vs. Athero+OI. Scale bars: 200 μm (top row in A and D ) and 50 μm (bottom row in A and D ).

Article Snippet: Immunohistochemical staining was performed to detect the macrophage marker Mac2 (sc-81728, Santa Cruz Biotechnology), and nuclei were counterstained with hematoxylin.

Techniques: Staining, Control, Comparison

Journal: Nature Communications

Article Title: The arginine methyltransferase PRMT7 promotes extravasation of monocytes resulting in tissue injury in COPD

doi: 10.1038/s41467-022-28809-4

Figure Lengend Snippet: a Heat map of the most significantly enriched gene lists in the lungs of COPD patients following gene set enrichment analysis (GSEA) of the GO molecular function set on publicly available array data from lung tissue (GSE76925) of healthy smokers ( n = 40) v COPD patients ( n = 111). Nominal P value generated by the GSEA software of the enrichment score relative to the null distribution shown. b Expression of genes involved in N -methyltransferase activity in COPD patients relative to healthy smokers taken from GSE76925, relative fold change, and P value calculated using the GEO2R interactive web tool running limma R. PRMT genes highlighted in red. c , d Expression of PRMT7 , in the lungs of healthy smokers and COPD patients, individuals are shown, expression relative to healthy smokers, taken from GSE76925 ( n = 40 smokers and n = 111 COPD patients) ( c ) and GSE27597 ( n = 16 smokers and n = 48 COPD patients) ( d ). e qPCR analysis of PRMT7 and TNF expression in human lung core biopsies from healthy controls ( n = 8) and COPD patients ( n = 18) relative to controls. f Western blot analysis of PRMT7 expression in lung core biopsies from healthy ( n = 17) and COPD patients ( n = 23), normalized to β-actin and shown relative to controls. g Representative images of immunofluorescence analysis for PRMT7 (Red) and the macrophage marker Galectin 3 (Green) in sections from core biopsies of healthy and COPD human lung ( n = 4, scale bar 20 μm) and sections from filtered air (FA) and cigarette smoke (CS) mouse lung ( n = 5, scale bar 25 μm). h Quantification of double-positive cells from ( g ). i , j mRNA expression levels of PRMT7 determined by qPCR in human monocytes isolated from blood of non-smokers ( n = 7) vs smokers ( n = 10) ( i ) and circulating monocytes ( n = 4 for FA and CS) ( i ) and alveolar macrophages ( n = 4 for FA and n = 3 for CS) ( j ) from B6 mice exposed to FA or CS for 3 days, relative to controls. k – m scRNA-Seq (Drop-Seq) analysis on the lungs of mice following exposure to FA ( n = 3) and CS for 2 ( n = 5) and 4 months ( n = 5). AM alveolar macrophages, CS-ind MØ CS-induced macrophages, IM interstitial macrophages, cMono classical monocytes. k Dot blot depicting Prmt7 expression (log-transformed, normalized UMI counts) and percentage of cells positive for Prmt7 in the myeloid cell compartment. l RNA velocity analysis of the myeloid compartment. (1) AM; (2) CS-ind MØ; (3) Lyve1 + /Cd163 + IM; (4) Lyve1−/Cd163− IM; (5) Prg4 + IM; (6) Ly6c2 + cMono; (7) Ly6c2- non-cMono; (8) Cd103 + /Clec9a + cDC; (9) Cd209 + /Cd11b + cDC; (10) Fscn1 + DC. m Fate probability mapping towards the CS-ind MØ population utilizing CellRank. n Heat map of gene expressions of PRMT6/7 , RAP1A , and ALOX5 in monocyte-like macrophages obtained from scRNA-Seq data of bronchoalveolar lavage from COPD patients ( n = 9) and healthy controls ( n = 6). Mean gene expression per donor is shown as a z-transformed value (across all donors). Data shown mean ± SD and individual patients or mice ( c – f and h – j ), P values shown in charts determined by two-tailed Mann–Whitney test ( c – f ), unpaired two-tailed Student’s t -test ( h – j ). Source data are provided as a Source Data file.

Article Snippet: After overnight incubation with a primary antibody against Galectin 3 (Rabbit anti-Galectin 3, 1:100, Cat. No. sc-20157, Santa Cruz), CD68 (Rabbit polyclonal, Cat. No. ab125212, 1:800, Abcam), ACSL4 (Mouse monoclonal, Cat.No. sc-365230, 1:50, Santa Cruz), ALOX5 (Mouse monoclonal, Cat.No. sc-136195, 1:50, Santa Cruz), and Pro-SPC (Rabbit polyclonal, Cat.No.

Techniques: Generated, Software, Expressing, Activity Assay, Western Blot, Immunofluorescence, Marker, Isolation, Dot Blot, Transformation Assay, Gene Expression, Two Tailed Test, MANN-WHITNEY

Journal: Nature Communications

Article Title: The arginine methyltransferase PRMT7 promotes extravasation of monocytes resulting in tissue injury in COPD

doi: 10.1038/s41467-022-28809-4

Figure Lengend Snippet: a Heat map of normalized enrichment score (NES) following GSEA of cell death pathway gene lists on array data from lung tissue of COPD patients (GSE47460-GPL14550; 145 COPD patients vs 91 healthy controls). b Expression of ferroptotic genes in the lungs of COPD patients relative to healthy controls taken from GSE47460-GPL14550, relative fold change and P value calculated using the GEO2R interactive web tool running limma R . c Western blot analysis of ACSL4 expression in lung core biopsies from healthy ( n = 3) and COPD patients ( n = 3), quantification relative to vinculin shown for individual patients. d Representative images of immunohistochemical analysis for ACSL4 (brown signal indicated by arrowhead, hematoxylin counterstained, scale bar 25 μm) in lung sections from COPD patients ( n = 4) and healthy controls ( n = 4). e Quantification of alveolar epithelial cells positive for ACSL4. f Heat map of NES following GSEA of cell death pathway gene lists on array data from mice exposed to 4 m chronic cigarette smoke (CS, n = 3) v filtered air (FA, n = 3). g Expression of ferroptotic genes in the lungs of mice exposed to 4 m chronic CS ( n = 3) relative to FA ( n = 3) taken from array data used in ( f ), relative fold change and P value calculated using the GEO2R interactive web tool running limma R. h , i Cells from whole lung suspensions of mice exposed to FA ( n = 3) or CS for 4 m ( n = 5), were analysed by scRNA-Seq (Drop-Seq). h UMAP of scRNA-Seq profiles (dots) colored by cell type. i UMAP plots showing expression of ACSL4 in scRNA-Seq profiles. j Representative images of immunohistochemical analysis for ACSL4 (red signal indicated by arrowheads, hematoxylin counterstained, scale bar 25 μm) in lung sections from WT and Prmt7 +/− mice exposed to FA or CS for 4 months ( n = 4–5 per group). k Quantification of alveolar epithelial cells positive for ACSL4 from ( j ) ( n = 5 for WT FA, Prmt7 +/ − FA, Prmt7 +/− CS and n = 4 for WT CS). l , m Mouse AT2 cells (MLE12) were treated with RSL3 (250 nM), or conditioned medium from control macrophages (MФ, RAW264.7 cells) or those polarized to a pro-inflammatory (IFNγ + LPS) or anti-inflammatory (IL4) phenotype for 48 h, representative data shown from a single experiment repeated twice. l Representative image showing PI uptake at 24 h (Scale bar 50 µm). m Number of dead AT2 cells at the time indicated, data shown from a single experiment repeated twice. n BODIPY oxidation in AT2 cells (MLE12) at 6 h after treatment with RSL3 (250 nM, n = 3) or conditioned medium from control macrophages ( n = 2) or those polarized to a pro-inflammatory ( n = 6) or anti-inflammatory ( n = 3) phenotype . o Number of dead AT2 cells (MLE12) by PI uptake at the time indicated following treatment with RSL3 (250 nM), or conditioned medium from macrophages (MФ, RAW264.7 cells) polarized to pro-inflammatory (IFNγ + LPS) plus 20 μM zVAD or 10 μM necrostatin-1s (Nec1-s), representative data shown from a single experiment repeated twice. p Western blot analysis of ACSL4 in AT2 cells (MLE12) treated for 24 h with conditioned medium from control macrophages ( n = 11) or those polarized to a pro-inflammatory ( n = 10) or anti-inflammatory ( n = 5) phenotype, normalized to β-actin and shown relative to control macrophages. q Western blot analysis of ALOX5 at 72 h in control macrophages ( n = 11) or those polarized to a pro-inflammatory ( n = 11) or anti-inflammatory ( n = 11) phenotype, normalized to β-actin and shown relative to control macrophages. r Western blot analysis of ACSL4 in AT2 cells (MLE12) treated for 24 h with conditioned medium from control macrophages or those polarized to a pro-inflammatory phenotype that were treated with siRNA against Alox5 , normalized to β-actin and shown relative to control macrophages, from individual experiments ( n = 3). Data shown mean ± SD, P values shown in charts determined by unpaired two-tailed Student’s t -test ( c , n ), two-tailed Mann–Whitney test ( e ), one-way ANOVA Bonferroni’s multiple comparisons test ( k , p , q ). Source data are provided as a Source Data file.

Article Snippet: After overnight incubation with a primary antibody against Galectin 3 (Rabbit anti-Galectin 3, 1:100, Cat. No. sc-20157, Santa Cruz), CD68 (Rabbit polyclonal, Cat. No. ab125212, 1:800, Abcam), ACSL4 (Mouse monoclonal, Cat.No. sc-365230, 1:50, Santa Cruz), ALOX5 (Mouse monoclonal, Cat.No. sc-136195, 1:50, Santa Cruz), and Pro-SPC (Rabbit polyclonal, Cat.No.

Techniques: Expressing, Western Blot, Immunohistochemical staining, Control, Two Tailed Test, MANN-WHITNEY