ccl28 cells  (Santa Cruz Biotechnology)

Journal: Nature Communications

Article Title: CCL28 contributes to angiogenesis and cardiac repair through CCR10 + endothelial cells after myocardial infarction in male mice

doi: 10.1038/s41467-025-64283-4

Figure Lengend Snippet: The hindlimb ischemia (HI) and myocardial ischemia (MI) were constructed on wild-type mice at specific time points (biological replicates, n = 4 per group). A The mRNA levels of Ccl27 and Ccl28 in the gastrocnemius of mice subjected to sham or HI surgery were determined by RT-qPCR at specific time points (biological replicates, n = 4 per group, One-Way ANOVA using Tukey multiple comparisons test). B The mRNA levels of Ccl27 and Ccl28 in the hearts of mice subjected to sham or MI surgery were determined by RT-qPCR at specific time points (biological replicates, n = 4 per group, One-Way ANOVA using Tukey multiple comparisons test). C Representative bands of CCL28 and ACTB protein in the gastrocnemius of mice subjected to sham or HI surgery measured by Western blot assay. Representative bands of CCL28 and ACTB protein in the hearts of mice subjected to sham or MI surgery measured by Western blot assay. D Quantification of CCL28 protein (biological replicates, n = 3 per group, One-Way ANOVA using Tukey multiple comparisons test). E Representative immunofluorescence images and quantification of CCL28 in gastrocnemius slice 7 days post HI and heart slice 3 days post MI with staining for CCL28 (red) and nuclei (blue). Scale bar, 200 μm. (biological replicates, n = 4 per group, One-Way ANOVA using Tukey multiple comparisons test). F Flow cytometry-based representative histogram (isotype control, red; CCL28 staining, blue) and quantification of CCL28 + cells isolated from the gastrocnemius of mice subjected to sham or HI surgery, and the hearts of mice subjected to sham or MI surgery at specific time points (biological replicates, n = 4 per group, One-Way ANOVA using Tukey multiple comparisons test). G Flow cytometry-based representative dot plots of CD11b + Ly6G - Ly6C - F4/80 + cells among the CCL28 + CD45 + cells 7 days post HI and 3 days post MI. H The percentage of multiple cells, including neutrophils (black), monocytes (yellow), macrophages (blue), and other cells (red), among the CCL28 + cells 7 days post HI and 3 days post MI (biological replicates, n = 4 per group). Quantitative real-time polymerase chain reaction, RT-qPCR; chemokine C-C motif ligand (CCL). Data are expressed as mean ± SD. Source data are provided as a Source Data file.

Article Snippet: After incubation with TruStain FcXTM antibody (101320, Biolegend) for 15 min to minimize non-specific binding, cells were then stained with the following antibodies for 30 min. For detection of CCL28 + cells: AF488 anti-CCL28(sc-376654, Santa Cruz), APC/FireTM 750 anti-CD45 (103154, Biolegend), PE/Cy7 anti-CD31(102524, Biolegend), APC anti-CD140a (135908, Biolegend), PE anti-Ly6C (128008, Biolegend), PerCP/Cyanine5.5 anti-Ly6G (127616, Biolegend), PE/Cy7 anti-CD11b (101216, Biolegend), APC anti-F4/80(123116, Biolegend).

Techniques: Construct, Quantitative RT-PCR, Western Blot, Immunofluorescence, Staining, Flow Cytometry, Control, Isolation, Real-time Polymerase Chain Reaction

Journal: Nature Communications

Article Title: CCL28 contributes to angiogenesis and cardiac repair through CCR10 + endothelial cells after myocardial infarction in male mice

doi: 10.1038/s41467-025-64283-4

Figure Lengend Snippet: Endothelial cells (ECs) are sorted by anti-CD31 magnetic beads and used for subsequent detection. A The mRNA levels of Ccr10 in ECs treated with mouse recombinant CCL28 protein (rCCL28) at different concentrations for 24 h were determined by RT-qPCR (biological replicates, n = 4 per group, One-Way ANOVA using Tukey multiple comparisons test). B Flow cytometry-based representative dot plots and quantification of CCR10 + ECs treated with rCCL28 at different concentrations (biological replicates, n = 4 per group, One-Way ANOVA using Tukey multiple comparisons test). C Gene set enrichment analysis (GSEA) for endothelial cells isolated from hearts of the Ccl28 -KO (KO) and littermate control mice (WT) mice subjected to MI surgery at 3 days, as tested by RNA-sequencing. D Representative bands of p-ERK1/2, ERK1/2, VEGFA, SOX5, CCR10 and ACTB protein measured by Western blot assay (biological replicates, n = 3 per group). E Representative bands of SOX5, CCR10 and ACTB protein measured by Western blot assay (biological replicates, n = 3 per group). F Western blot analysis of input proteins and proteins immunoprecipitated with either IgG or SOX5 antibody. G ChIP analysis of the Ccr10 promoter site 3 (−400 to +200) in ECs by RT-qPCR (biological replicates, n = 3 per group, Unpaired two-tailed Student’s t -test). H Dual-luciferase reporter assay was performed after co-transfection with a reporter vector in NIH/3T3 (biological replicates, n = 4 per group, Two-Way ANOVA using Tukey multiple comparisons test). I , J The sprouting ability (biological replicates, n = 4 per group, Unpaired two-tailed Student’s t -test. scale bar, 100 μm) and K aging proportion (biological replicates, n = 4 per group, Unpaired two-tailed Student’s t -test. scale bar, 250 μm) of ECs were measured. L Mechanism diagram of CCL28-induced CCR10/ERK/SOX5 positive feedback signaling. Data are expressed as mean ± SD. Source data are provided as a Source Data file.

Article Snippet: After incubation with TruStain FcXTM antibody (101320, Biolegend) for 15 min to minimize non-specific binding, cells were then stained with the following antibodies for 30 min. For detection of CCL28 + cells: AF488 anti-CCL28(sc-376654, Santa Cruz), APC/FireTM 750 anti-CD45 (103154, Biolegend), PE/Cy7 anti-CD31(102524, Biolegend), APC anti-CD140a (135908, Biolegend), PE anti-Ly6C (128008, Biolegend), PerCP/Cyanine5.5 anti-Ly6G (127616, Biolegend), PE/Cy7 anti-CD11b (101216, Biolegend), APC anti-F4/80(123116, Biolegend).

Techniques: Magnetic Beads, Recombinant, Quantitative RT-PCR, Flow Cytometry, Isolation, Control, RNA Sequencing, Western Blot, Immunoprecipitation, Two Tailed Test, Luciferase, Reporter Assay, Cotransfection, Plasmid Preparation

Journal: Nature Communications

Article Title: CCL28 contributes to angiogenesis and cardiac repair through CCR10 + endothelial cells after myocardial infarction in male mice

doi: 10.1038/s41467-025-64283-4

Figure Lengend Snippet: The hindlimb ischemia (HI) and myocardial ischemia (MI) were constructed on Ccl28- KO (KO) and littermate control mice (WT). A Representative images and quantification of blood flow in mice examined by laser Doppler imaging at specific time points (biological replicates, n = 8 per group, Two-Way ANOVA using Tukey multiple comparisons test). B Representative immunofluorescence images and quantification of vessel numbers in the WT and Ccl28 -KO mice of gastrocnemius 7 days post HI with staining for CD31 (red) and nuclei (blue) (biological replicates, n = 5 per group, Unpaired two-tailed Student’s t -test). Scale bar, 200 μm. C The percentage of collagen volume fraction in the WT and Ccl28 -KO mice of gastrocnemius 14 days post HI was assessed by Masson’s trichrome staining (biological replicates, n = 5 per group, Unpaired two-tailed Student’s t -test). Scale bar, 100 μm. D Representative immunofluorescence images and quantification of vessel numbers in the WT and Ccl28 -KO of hearts 7 days post MI with staining for CD31 (red) and nuclei (blue) (biological replicates, n = 5 per group, Unpaired two-tailed Student’s t -test). Scale bar, 200 μm. E , F The percentage of infarct size and collagen volume fraction in the WT and Ccl28 -KO of hearts 7 days post MI were assessed by Masson’s trichrome staining (biological replicates, n = 4 per group, Unpaired two-tailed Student’s t -test). Scale bar, 1 mm. G Echocardiography was performed to assess cardiac function of the left ventricle in the WT and Ccl28 -KO mice subjected to sham or MI surgery at specific time points (biological replicates, n = 6 per group, Two-Way ANOVA using Tukey multiple comparisons test). H The FITC-Dextran (green) penetrance was used to detect vascular leakage in the WT and Ccl28 -KO of hearts 3 days post MI (biological replicates, n = 6 per group, Unpaired two-tailed Student’s t -test). Scale bar, 100 μm. Data are expressed as mean ± SD. Source data are provided as a Source Data file.

Article Snippet: After incubation with TruStain FcXTM antibody (101320, Biolegend) for 15 min to minimize non-specific binding, cells were then stained with the following antibodies for 30 min. For detection of CCL28 + cells: AF488 anti-CCL28(sc-376654, Santa Cruz), APC/FireTM 750 anti-CD45 (103154, Biolegend), PE/Cy7 anti-CD31(102524, Biolegend), APC anti-CD140a (135908, Biolegend), PE anti-Ly6C (128008, Biolegend), PerCP/Cyanine5.5 anti-Ly6G (127616, Biolegend), PE/Cy7 anti-CD11b (101216, Biolegend), APC anti-F4/80(123116, Biolegend).

Techniques: Construct, Control, Imaging, Immunofluorescence, Staining, Two Tailed Test

Journal: Nature Communications

Article Title: CCL28 contributes to angiogenesis and cardiac repair through CCR10 + endothelial cells after myocardial infarction in male mice

doi: 10.1038/s41467-025-64283-4

Figure Lengend Snippet: The hindlimb ischemia (HI) and myocardial ischemia (MI) were constructed on wild-type mice treated with mouse recombinant CCL28 protein (rCCL28, 50 μg/kg, every four days) or vehicle. A Representative images and quantification of blood flow in mice examined by laser Doppler imaging at specific time points (biological replicates, biological replicates, n = 8 per group, Two-Way ANOVA using Tukey multiple comparisons test). B Representative immunofluorescence images and quantification of vessel numbers in the Vehicle and rCCL28 groups of gastrocnemius 7 days post HI with staining for CD31 (red) and nuclei (blue) (biological replicates, n = 5 per group, Unpaired two-tailed Student’s t -test). Scale bar, 200 μm. C The percentage of collagen volume fraction in the Vehicle and rCCL28 groups mice of gastrocnemius 14 days post HI was assessed by Masson’s trichrome staining (biological replicates, n = 5 per group, Unpaired two-tailed Student’s t -test). Scale bar, 100 μm. D Representative immunofluorescence images and quantification of vessel numbers in the Vehicle and rCCL28 groups of hearts 3 days post MI with staining for CD31 (red) and nuclei (blue) (biological replicates, n = 5 per group, Unpaired two-tailed Student’s t -test). Scale bar, 200 μm. E , F The percentage of infarct size and collagen volume fraction in the Vehicle and rCCL28 groups of hearts 7 days post MI were assessed by Masson’s trichrome staining (biological replicates, n = 4 per group, Unpaired two-tailed Student’s t -test). Scale bar, 1 mm. G Echocardiography was performed to assess cardiac function of the left ventricle in the Vehicle and rCCL28 groups subjected to sham or MI surgery at specific time points (biological replicates, n = 6 per group, Two-Way ANOVA using Tukey multiple comparisons test). H The FITC-Dextran (green) penetrance was used to detect vascular leakage in the Vehicle and rCCL28 groups of hearts 3 days post MI (biological replicates, n = 6 per group, Unpaired two-tailed Student’s t -test). Scale bar, 100 μm. Data are expressed as mean ± SD. Source data are provided as a Source Data file.

Article Snippet: After incubation with TruStain FcXTM antibody (101320, Biolegend) for 15 min to minimize non-specific binding, cells were then stained with the following antibodies for 30 min. For detection of CCL28 + cells: AF488 anti-CCL28(sc-376654, Santa Cruz), APC/FireTM 750 anti-CD45 (103154, Biolegend), PE/Cy7 anti-CD31(102524, Biolegend), APC anti-CD140a (135908, Biolegend), PE anti-Ly6C (128008, Biolegend), PerCP/Cyanine5.5 anti-Ly6G (127616, Biolegend), PE/Cy7 anti-CD11b (101216, Biolegend), APC anti-F4/80(123116, Biolegend).

Techniques: Construct, Recombinant, Imaging, Immunofluorescence, Staining, Two Tailed Test

Journal: Nature Communications

Article Title: CCL28 contributes to angiogenesis and cardiac repair through CCR10 + endothelial cells after myocardial infarction in male mice

doi: 10.1038/s41467-025-64283-4

Figure Lengend Snippet: A – H the hindlimb ischemia (HI) and myocardial ischemia (MI) were constructed on wild-type and db/db mice. A Representative bands of CCL28 and ACTB protein in the gastrocnemius of mice subjected to sham or HI surgery at 7 days was measured by Western blot assay (biological replicates, n = 3 per group). B Representative bands of CCL28 and ACTB protein in the hearts of mice subjected to sham or MI surgery at 3 days measured by Western blot assay (biological replicates, n = 3 per group). C Representative bands of CCR10 and ACTB protein in the gastrocnemius of mice subjected to sham or HI surgery at 7 days measured by Western blot assay (biological replicates, n = 3 per group). D Representative bands of CCR10 and ACTB protein in the hearts of mice subjected to sham or MI surgery at 3 days measured by Western blot assay (biological replicates, n = 3 per group). E – H Flow cytometry-based representative dot plots and quantification of CCR10 + CD31 + endothelial cells isolated from the gastrocnemius of mice subjected to sham or HI surgery at 7 days, and the hearts of mice subjected to sham or MI surgery at 3 days (biological replicates, n = 4 per group, Unpaired two-tailed Student’s t -test). I – M the hindlimb ischemia (HI) and myocardial ischemia (MI) were constructed on db/db mice treated with mouse recombinant CCL28 protein (rCCL28, 50 μg/kg, every four days) or vehicle. I – L Representative immunofluorescence images and quantification of vessel numbers in the Vehicle and rCCL28 groups of gastrocnemius 7 days post HI, and hearts 3 days post MI with staining for CD31 (red) and nuclei (blue) (biological replicates, n = 5 per group, Unpaired two-tailed Student’s t -test). Scale bar, 200 μm. M Representative images and quantification of blood flow in mice examined by laser Doppler imaging at specific time points (biological replicates, n = 8 per group, Two-Way ANOVA using Tukey multiple comparisons test). Data are expressed as mean ± SD. Source data are provided as a Source Data file.

Article Snippet: After incubation with TruStain FcXTM antibody (101320, Biolegend) for 15 min to minimize non-specific binding, cells were then stained with the following antibodies for 30 min. For detection of CCL28 + cells: AF488 anti-CCL28(sc-376654, Santa Cruz), APC/FireTM 750 anti-CD45 (103154, Biolegend), PE/Cy7 anti-CD31(102524, Biolegend), APC anti-CD140a (135908, Biolegend), PE anti-Ly6C (128008, Biolegend), PerCP/Cyanine5.5 anti-Ly6G (127616, Biolegend), PE/Cy7 anti-CD11b (101216, Biolegend), APC anti-F4/80(123116, Biolegend).

Techniques: Construct, Western Blot, Flow Cytometry, Isolation, Two Tailed Test, Recombinant, Immunofluorescence, Staining, Imaging

Journal: Nature Communications

Article Title: CCL28 contributes to angiogenesis and cardiac repair through CCR10 + endothelial cells after myocardial infarction in male mice

doi: 10.1038/s41467-025-64283-4

Figure Lengend Snippet: A Flowchart of study participants. B The serum CCL28 levels were assessed by ELISA assay (biological replicates, the numbers of participants with Rentrop score 0, 1, 2, 3 were 24, 133, 145, and 114, respectively. Kruskal-Wallis test using Dunn multiple comparisons test). In detail, the boxplots depict the distribution of CCL28 levels across four Rentrop classifications (Rentrop 0–Rentrop 3). Each box represents the interquartile range (IQR), with the lower boundary corresponding to the 25th percentile (Q1) and the upper boundary to the 75th percentile (Q3); the median is indicated by a central line. The whiskers extend to the 10th and 90th percentiles, respectively, capturing the central 80% of the data, while the minimum and maximum values within each Rentrop group are explicitly marked to illustrate the full data range. Specifically, for Rentrop0, the median is 499.5 (Q1: 493.25, Q3: 561.5) pg/mL, with whiskers spanning from 447.5 (10th percentile) to 738.5 (90th percentile) pg/mL, and the minimum and maximum values are 408 and 807 pg/mL, respectively; for Rentrop1, the median is 592 (Q1: 545, Q3: 686) pg/mL, with whiskers from 507.8 to 833.2 pg/mL, and the minimum and maximum values are 411 and 1832 pg/mL; for Rentrop2, the median is 611 (Q1: 567, Q3: 746) pg/mL, with whiskers from 534.6 to 1010 pg/mL, and the minimum and maximum values are 514 and 2519 pg/mL; and for Rentrop3, the median is 679.5 (Q1: 576.25, Q3: 826.75) pg/mL, with whiskers from 545.5 to 1162.5 pg/mL, and the minimum and maximum values are 408 and 2230 pg/mL. C Receiver operating characteristic curve analysis of CCL28 for predicting CCV (red). D Receiver operating characteristic curve analysis of CTO risk model (red) and its modifications (blue) for predicting CCV. E Graphical abstract of main findings. Source data are provided as a Source Data file.

Article Snippet: After incubation with TruStain FcXTM antibody (101320, Biolegend) for 15 min to minimize non-specific binding, cells were then stained with the following antibodies for 30 min. For detection of CCL28 + cells: AF488 anti-CCL28(sc-376654, Santa Cruz), APC/FireTM 750 anti-CD45 (103154, Biolegend), PE/Cy7 anti-CD31(102524, Biolegend), APC anti-CD140a (135908, Biolegend), PE anti-Ly6C (128008, Biolegend), PerCP/Cyanine5.5 anti-Ly6G (127616, Biolegend), PE/Cy7 anti-CD11b (101216, Biolegend), APC anti-F4/80(123116, Biolegend).

Techniques: Enzyme-linked Immunosorbent Assay