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Expression of <t>ADAM8</t> in human and mice (A) Dot heatmap of ADAM family expression profiles in the GSE97320 dataset, which includes gene expression profiles from the peripheral blood of healthy people (n = 3) and AMI patients (n = 3). Horizontal ordinate means represent the adjusted P value, and ADAM8 has the lowest P value. (B) Heatmap of ADAM family expression profiles in the GSE97320 dataset. (C) Different cell clusters from heart of sham and MI mice based on bioinformatics of GSE216211 dataset. (D) ADAM8 expression in different cell clusters from sham and MI mice based on bioinformatics of GSE216211 dataset. (E) Expression of ADAM8 in macrophages from baseline mice, and 3 and 7 days after coronary occlusion in GSE187701 dataset based on FPKM from GEO datasets of PubMed (p < 0.001, One-way ANOVA and Tukey's test). (F) Expression of ADAM8 in plasma of control and AMI patients (p = 0.0169, Mann Whitney test) (Left). Expression of ADAM8 in plasma of Non-MI, STEMI and NSTEMI patients (p = 0.037, One-way ANOVA and SNK test) (Right). (G) Western blot of ADAM8 expression in hearts of mice with sham or MI operation after day 3 and day 7 (Left). Protein quantification of ADAM8 (Right) (n = 6) (p < 0.0001, Welch's ANOVA test). (H) Relative mRNA of ADAM8 in hearts of mice with sham or MI operation after day 3 and day 7 (n = 5) (p < 0.001, Kruskal-Wallis test). (I-J) Representative immunofluorescence images of ADAM8 (Red) and F4/80 (Green) or Ly6G (Green) in heart sections from WT mice receiving sham operation or infarcted areas from mice receiving MI operation after day 3 and day 7. n = 3, Scale bar = 50 μm. *p < 0.05, **p < 0.01, ***p < 0.001. “n value” refers to the number of biological replicates. ADAM8: A disintegrin and metallopeptidase domain 8. AMI: Acute myocardial infarction. FPKM: Fragments Per Kilobase Million.
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Prognostic and immune cell correlation analysis of PRDEGs. a – j KM curves were independently analyzed at the transcriptome level to determine the prognostic value of these PRDEGs ( a – j : HEYL, FSTL3, HOMER3, CALB2, FABP4, MEGF6, <t>ADAM8,</t> TIMP1, CLCA1, NOS2). k The results of the correlation analysis between the PRDEGs and each infiltrating immunocyte are displayed in a heatmap
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Prognostic and immune cell correlation analysis of PRDEGs. a – j KM curves were independently analyzed at the transcriptome level to determine the prognostic value of these PRDEGs ( a – j : HEYL, FSTL3, <t>HOMER3,</t> CALB2, FABP4, MEGF6, ADAM8, TIMP1, CLCA1, NOS2). k The results of the correlation analysis between the PRDEGs and each infiltrating immunocyte are displayed in a heatmap
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Expression of ADAM8 in human and mice (A) Dot heatmap of ADAM family expression profiles in the GSE97320 dataset, which includes gene expression profiles from the peripheral blood of healthy people (n = 3) and AMI patients (n = 3). Horizontal ordinate means represent the adjusted P value, and ADAM8 has the lowest P value. (B) Heatmap of ADAM family expression profiles in the GSE97320 dataset. (C) Different cell clusters from heart of sham and MI mice based on bioinformatics of GSE216211 dataset. (D) ADAM8 expression in different cell clusters from sham and MI mice based on bioinformatics of GSE216211 dataset. (E) Expression of ADAM8 in macrophages from baseline mice, and 3 and 7 days after coronary occlusion in GSE187701 dataset based on FPKM from GEO datasets of PubMed (p < 0.001, One-way ANOVA and Tukey's test). (F) Expression of ADAM8 in plasma of control and AMI patients (p = 0.0169, Mann Whitney test) (Left). Expression of ADAM8 in plasma of Non-MI, STEMI and NSTEMI patients (p = 0.037, One-way ANOVA and SNK test) (Right). (G) Western blot of ADAM8 expression in hearts of mice with sham or MI operation after day 3 and day 7 (Left). Protein quantification of ADAM8 (Right) (n = 6) (p < 0.0001, Welch's ANOVA test). (H) Relative mRNA of ADAM8 in hearts of mice with sham or MI operation after day 3 and day 7 (n = 5) (p < 0.001, Kruskal-Wallis test). (I-J) Representative immunofluorescence images of ADAM8 (Red) and F4/80 (Green) or Ly6G (Green) in heart sections from WT mice receiving sham operation or infarcted areas from mice receiving MI operation after day 3 and day 7. n = 3, Scale bar = 50 μm. *p < 0.05, **p < 0.01, ***p < 0.001. “n value” refers to the number of biological replicates. ADAM8: A disintegrin and metallopeptidase domain 8. AMI: Acute myocardial infarction. FPKM: Fragments Per Kilobase Million.

Journal: Journal of Advanced Research

Article Title: ADAM8 deficiency in macrophages promotes cardiac repair after myocardial infarction via ANXA2-mTOR-autophagy pathway

doi: 10.1016/j.jare.2024.07.037

Figure Lengend Snippet: Expression of ADAM8 in human and mice (A) Dot heatmap of ADAM family expression profiles in the GSE97320 dataset, which includes gene expression profiles from the peripheral blood of healthy people (n = 3) and AMI patients (n = 3). Horizontal ordinate means represent the adjusted P value, and ADAM8 has the lowest P value. (B) Heatmap of ADAM family expression profiles in the GSE97320 dataset. (C) Different cell clusters from heart of sham and MI mice based on bioinformatics of GSE216211 dataset. (D) ADAM8 expression in different cell clusters from sham and MI mice based on bioinformatics of GSE216211 dataset. (E) Expression of ADAM8 in macrophages from baseline mice, and 3 and 7 days after coronary occlusion in GSE187701 dataset based on FPKM from GEO datasets of PubMed (p < 0.001, One-way ANOVA and Tukey's test). (F) Expression of ADAM8 in plasma of control and AMI patients (p = 0.0169, Mann Whitney test) (Left). Expression of ADAM8 in plasma of Non-MI, STEMI and NSTEMI patients (p = 0.037, One-way ANOVA and SNK test) (Right). (G) Western blot of ADAM8 expression in hearts of mice with sham or MI operation after day 3 and day 7 (Left). Protein quantification of ADAM8 (Right) (n = 6) (p < 0.0001, Welch's ANOVA test). (H) Relative mRNA of ADAM8 in hearts of mice with sham or MI operation after day 3 and day 7 (n = 5) (p < 0.001, Kruskal-Wallis test). (I-J) Representative immunofluorescence images of ADAM8 (Red) and F4/80 (Green) or Ly6G (Green) in heart sections from WT mice receiving sham operation or infarcted areas from mice receiving MI operation after day 3 and day 7. n = 3, Scale bar = 50 μm. *p < 0.05, **p < 0.01, ***p < 0.001. “n value” refers to the number of biological replicates. ADAM8: A disintegrin and metallopeptidase domain 8. AMI: Acute myocardial infarction. FPKM: Fragments Per Kilobase Million.

Article Snippet: Immunoprecipitation was also performed with the anti-ADAM8 antibody (IP: ADAM8) or IgG (IP: IgG) and RAW264.7 cells, and the protein lysates were used for LC-MS/MS (Applied Protein Technology Co. Ltd, Shanghai).

Techniques: Expressing, Gene Expression, Clinical Proteomics, Control, MANN-WHITNEY, Western Blot, Immunofluorescence

Macrophage ADAM8 specific knockout improved heart function and survival of AMI mice (A) Western blot validation of ADAM8 KO BMDM (p = 0.0033, Unpaired t test with Welch's correction) (n = 8). (B) qRT-PCR validation of ADAM8 KO in BMDM (p = 0.0025, unpaired t test with Welch's correction) (n = 3). (C) Echocardiogram and related statistics of Flox and KO mice (n = 8). LVEF, p < 0.001; FS, p < 0.001; LVID;d, p = 0.040; LVID;s, p = 0.0035, unpaired t test. (D) HW/BW (p = 0.012, unpaired t test) and LW/BW (p = 0.0002, Mann Whitney test) in Flox and KO mice with MI (n = 9). (E) Western blot of VEGFA (p = 0.027, unpaired t test with Welch's correction), collagen I (p = 0.034, unpaired t test) and collagen III (p = 0.004, unpaired t test with Welch's correction) expression in hearts of Flox and KO mice after MI (n = 9). (F) Representative immunofluorescence images of CD31 in Flox and KO mice (n = 7), Scale bar = 50 μm (p = 0.001), unpaired t test. (G) Representative Masson images of Flox and KO mice (n = 8–9), Scale bar=1 mm (p = 0.001), unpaired t test. (H) Cumulative mortality within 28 days after MI in Flox and KO mice (n = 30) (p = 0.041, Logrank test). (I-K) 4D label-free proteomics based on DIA in flox and KO mice with sham or MI operation after 14 days. *p < 0.05, **p < 0.01, ***p < 0.001. “n value” refers to the number of biological replicates. ADAM8: A disintegrin and metallopeptidase domain 8. KO: Knockout. HW/BW: Heart weight/Body weight. LW/BW: Lung weight/ Body weight. MI: Myocardial infarction. DIA: Data-independent Acquisition.

Journal: Journal of Advanced Research

Article Title: ADAM8 deficiency in macrophages promotes cardiac repair after myocardial infarction via ANXA2-mTOR-autophagy pathway

doi: 10.1016/j.jare.2024.07.037

Figure Lengend Snippet: Macrophage ADAM8 specific knockout improved heart function and survival of AMI mice (A) Western blot validation of ADAM8 KO BMDM (p = 0.0033, Unpaired t test with Welch's correction) (n = 8). (B) qRT-PCR validation of ADAM8 KO in BMDM (p = 0.0025, unpaired t test with Welch's correction) (n = 3). (C) Echocardiogram and related statistics of Flox and KO mice (n = 8). LVEF, p < 0.001; FS, p < 0.001; LVID;d, p = 0.040; LVID;s, p = 0.0035, unpaired t test. (D) HW/BW (p = 0.012, unpaired t test) and LW/BW (p = 0.0002, Mann Whitney test) in Flox and KO mice with MI (n = 9). (E) Western blot of VEGFA (p = 0.027, unpaired t test with Welch's correction), collagen I (p = 0.034, unpaired t test) and collagen III (p = 0.004, unpaired t test with Welch's correction) expression in hearts of Flox and KO mice after MI (n = 9). (F) Representative immunofluorescence images of CD31 in Flox and KO mice (n = 7), Scale bar = 50 μm (p = 0.001), unpaired t test. (G) Representative Masson images of Flox and KO mice (n = 8–9), Scale bar=1 mm (p = 0.001), unpaired t test. (H) Cumulative mortality within 28 days after MI in Flox and KO mice (n = 30) (p = 0.041, Logrank test). (I-K) 4D label-free proteomics based on DIA in flox and KO mice with sham or MI operation after 14 days. *p < 0.05, **p < 0.01, ***p < 0.001. “n value” refers to the number of biological replicates. ADAM8: A disintegrin and metallopeptidase domain 8. KO: Knockout. HW/BW: Heart weight/Body weight. LW/BW: Lung weight/ Body weight. MI: Myocardial infarction. DIA: Data-independent Acquisition.

Article Snippet: Immunoprecipitation was also performed with the anti-ADAM8 antibody (IP: ADAM8) or IgG (IP: IgG) and RAW264.7 cells, and the protein lysates were used for LC-MS/MS (Applied Protein Technology Co. Ltd, Shanghai).

Techniques: Knock-Out, Western Blot, Biomarker Discovery, Quantitative RT-PCR, MANN-WHITNEY, Expressing, Immunofluorescence, Data-independent acquisition

Overexpression of ADAM8 can exacerbate myocardial dysfunction improved by macrophage-specific ADAM8 loss (A) Echocardiogram in KO+NC and KO+OV mice (n = 6). LVEF, p = 0.013; FS, p = 0.014, unpaired t test. (B) HW/BW (p = 0.026, Mann Whitney test) and LW/BW (p = 0.013, unpaired t test) in KO+NC and KO+OV mice with MI (n = 6). (C) Western blot of VEGFA (p = 0.033), collagen I (p < 0.001) and collagen III (p = 0.008) expression in hearts of KO+NC and KO+OV mice after MI (n = 6), unpaired t test. (D) Representative immunofluorescence images of CD31 in KO+NC and KO+OV mice (n = 5–6), Scale bar=50 μm (p = 0.038, unpaired t test with Welch's correction). (E) Representative Masson images of KO+NC and KO+OV mice (n = 5–6), Scale bar = 1 mm (p = 0.007, unpaired t test). (F) Cumulative mortality within 28 days after MI in KO+NC (n = 15) and KO+OV mice (n = 16) (p = 0.044, Logrank test). *p < 0.05, **p < 0.01, ***p < 0.001. “n value” refers to the number of biological replicates. ADAM8: A disintegrin and metallopeptidase domain 8. KO: Knockout. OV: Overexpression. HW/BW: Heart weight/Body weight. LW/BW: Lung weight/ Body weight. MI: Myocardial infarction.

Journal: Journal of Advanced Research

Article Title: ADAM8 deficiency in macrophages promotes cardiac repair after myocardial infarction via ANXA2-mTOR-autophagy pathway

doi: 10.1016/j.jare.2024.07.037

Figure Lengend Snippet: Overexpression of ADAM8 can exacerbate myocardial dysfunction improved by macrophage-specific ADAM8 loss (A) Echocardiogram in KO+NC and KO+OV mice (n = 6). LVEF, p = 0.013; FS, p = 0.014, unpaired t test. (B) HW/BW (p = 0.026, Mann Whitney test) and LW/BW (p = 0.013, unpaired t test) in KO+NC and KO+OV mice with MI (n = 6). (C) Western blot of VEGFA (p = 0.033), collagen I (p < 0.001) and collagen III (p = 0.008) expression in hearts of KO+NC and KO+OV mice after MI (n = 6), unpaired t test. (D) Representative immunofluorescence images of CD31 in KO+NC and KO+OV mice (n = 5–6), Scale bar=50 μm (p = 0.038, unpaired t test with Welch's correction). (E) Representative Masson images of KO+NC and KO+OV mice (n = 5–6), Scale bar = 1 mm (p = 0.007, unpaired t test). (F) Cumulative mortality within 28 days after MI in KO+NC (n = 15) and KO+OV mice (n = 16) (p = 0.044, Logrank test). *p < 0.05, **p < 0.01, ***p < 0.001. “n value” refers to the number of biological replicates. ADAM8: A disintegrin and metallopeptidase domain 8. KO: Knockout. OV: Overexpression. HW/BW: Heart weight/Body weight. LW/BW: Lung weight/ Body weight. MI: Myocardial infarction.

Article Snippet: Immunoprecipitation was also performed with the anti-ADAM8 antibody (IP: ADAM8) or IgG (IP: IgG) and RAW264.7 cells, and the protein lysates were used for LC-MS/MS (Applied Protein Technology Co. Ltd, Shanghai).

Techniques: Over Expression, MANN-WHITNEY, Western Blot, Expressing, Immunofluorescence, Knock-Out

Bone marrow transplantation validates the adverse role of ADAM8 in myocardial infarction (A) Schematic diagram of bone marrow transplantation. (B) Echocardiogram in Flox-KO and KO-KO mice (n = 7–8). LVEF, p < 0.001; FS, p < 0.001; LVID;d, p = 0.021;LVID;s, p = 0.004, unpaired t test. (C) HW/BW (p = 0.021) and LW/BW (p = 0.058) in Flox-KO and KO-KO mice with MI (n = 7), unpaired t test. (D) Western blot of VEGFA (p = 0.002, Mann Whitney test), collagen I (p = 0.004) and collagen III (p = 0.013) expression in hearts of Flox-KO and KO-KO mice after MI (n = 6), unpaired t test. (E) Representative immunofluorescence images of CD31 in Flox-KO and KO-KO mice (n = 7–8), Scale bar=50 μm (p = 0.002, unpaired t test with Welch's correction). (F) Representative Masson images of Flox-KO and KO-KO mice (n = 7–8), Scale bar = 1 mm (p = 0.008, unpaired t test with Welch's correction). (G) Cumulative mortality within 28 days after MI in Flox-KO (n = 18) and KO-KO mice (n = 17) (p = 0.024, Logrank test). *p < 0.05, **p < 0.01, ***p < 0.001. “n value” refers to the number of biological replicates. ADAM8: A disintegrin and metallopeptidase domain 8. KO: Knockout. HW/BW: Heart weight/Body weight. LW/BW: Lung weight/ Body weight. MI: Myocardial infarction.

Journal: Journal of Advanced Research

Article Title: ADAM8 deficiency in macrophages promotes cardiac repair after myocardial infarction via ANXA2-mTOR-autophagy pathway

doi: 10.1016/j.jare.2024.07.037

Figure Lengend Snippet: Bone marrow transplantation validates the adverse role of ADAM8 in myocardial infarction (A) Schematic diagram of bone marrow transplantation. (B) Echocardiogram in Flox-KO and KO-KO mice (n = 7–8). LVEF, p < 0.001; FS, p < 0.001; LVID;d, p = 0.021;LVID;s, p = 0.004, unpaired t test. (C) HW/BW (p = 0.021) and LW/BW (p = 0.058) in Flox-KO and KO-KO mice with MI (n = 7), unpaired t test. (D) Western blot of VEGFA (p = 0.002, Mann Whitney test), collagen I (p = 0.004) and collagen III (p = 0.013) expression in hearts of Flox-KO and KO-KO mice after MI (n = 6), unpaired t test. (E) Representative immunofluorescence images of CD31 in Flox-KO and KO-KO mice (n = 7–8), Scale bar=50 μm (p = 0.002, unpaired t test with Welch's correction). (F) Representative Masson images of Flox-KO and KO-KO mice (n = 7–8), Scale bar = 1 mm (p = 0.008, unpaired t test with Welch's correction). (G) Cumulative mortality within 28 days after MI in Flox-KO (n = 18) and KO-KO mice (n = 17) (p = 0.024, Logrank test). *p < 0.05, **p < 0.01, ***p < 0.001. “n value” refers to the number of biological replicates. ADAM8: A disintegrin and metallopeptidase domain 8. KO: Knockout. HW/BW: Heart weight/Body weight. LW/BW: Lung weight/ Body weight. MI: Myocardial infarction.

Article Snippet: Immunoprecipitation was also performed with the anti-ADAM8 antibody (IP: ADAM8) or IgG (IP: IgG) and RAW264.7 cells, and the protein lysates were used for LC-MS/MS (Applied Protein Technology Co. Ltd, Shanghai).

Techniques: Transplantation Assay, Western Blot, MANN-WHITNEY, Expressing, Immunofluorescence, Knock-Out

Bevacizumab aggravates the cardiac function of ADAM8 KO mice after MI (A) Echocardiogram in KO and KO + Be mice (n = 7–8). LVEF, p = 0.018; FS, p = 0.019; LVID;d, p = 0.008;LVID;s, p = 0.007, unpaired t test. (B) HW/BW (p = 0.018, unpaired t test) and LW/BW (p = 0.001, Mann Whitney test) in KO and KO + Be mice with MI (n = 7–8). (C) Western blot of collagen I (p = 0.005, Mann Whitney test), collagen III (p = 0.005, unpaired t test with Welch’s correction), and VEGFA (p = 0.006, unpaired t test) expression in hearts of KO and KO + Be mice after MI (n = 6). (D) Representative immunofluorescence images of CD31 in KO and KO + Be mice (n = 6), Scale bar=50 μm (p = 0.0001, unpaired t test). (E) Representative Masson images of KO and KO + Be mice (n = 5), Scale bar = 1 mm (p = 0.023, unpaired t test). (F) Cumulative mortality within 28 days after MI in KO and KO + Be mice (n = 11) (p = 0.031, Logrank test). *p < 0.05, **p < 0.01, ***p < 0.001. “n value” refers to the number of biological replicates. ADAM8: A disintegrin and metallopeptidase domain 8. Be: Bevacizumab. KO: Knockout. HW/BW: Heart weight/Body weight. LW/BW: Lung weight/ Body weight. MI: Myocardial infarction.

Journal: Journal of Advanced Research

Article Title: ADAM8 deficiency in macrophages promotes cardiac repair after myocardial infarction via ANXA2-mTOR-autophagy pathway

doi: 10.1016/j.jare.2024.07.037

Figure Lengend Snippet: Bevacizumab aggravates the cardiac function of ADAM8 KO mice after MI (A) Echocardiogram in KO and KO + Be mice (n = 7–8). LVEF, p = 0.018; FS, p = 0.019; LVID;d, p = 0.008;LVID;s, p = 0.007, unpaired t test. (B) HW/BW (p = 0.018, unpaired t test) and LW/BW (p = 0.001, Mann Whitney test) in KO and KO + Be mice with MI (n = 7–8). (C) Western blot of collagen I (p = 0.005, Mann Whitney test), collagen III (p = 0.005, unpaired t test with Welch’s correction), and VEGFA (p = 0.006, unpaired t test) expression in hearts of KO and KO + Be mice after MI (n = 6). (D) Representative immunofluorescence images of CD31 in KO and KO + Be mice (n = 6), Scale bar=50 μm (p = 0.0001, unpaired t test). (E) Representative Masson images of KO and KO + Be mice (n = 5), Scale bar = 1 mm (p = 0.023, unpaired t test). (F) Cumulative mortality within 28 days after MI in KO and KO + Be mice (n = 11) (p = 0.031, Logrank test). *p < 0.05, **p < 0.01, ***p < 0.001. “n value” refers to the number of biological replicates. ADAM8: A disintegrin and metallopeptidase domain 8. Be: Bevacizumab. KO: Knockout. HW/BW: Heart weight/Body weight. LW/BW: Lung weight/ Body weight. MI: Myocardial infarction.

Article Snippet: Immunoprecipitation was also performed with the anti-ADAM8 antibody (IP: ADAM8) or IgG (IP: IgG) and RAW264.7 cells, and the protein lysates were used for LC-MS/MS (Applied Protein Technology Co. Ltd, Shanghai).

Techniques: MANN-WHITNEY, Western Blot, Expressing, Immunofluorescence, Knock-Out

ADAM8 regulates VEGFA secretion and inflammation in macrophages via autophagy pathway (A) Western blot of IL-1β, IL-18, TNF-α and VEGFA in Flox and KO BMDMs treated with or without LPS (1 μg/mL for 12 h) and IL-4 (20 ng/mL for 12 h) (n = 3–4), unpaired t test among different treatment groups. (B) Levels of IL-1β, IL-18, TNF-α and VEGFA in supernatants of Flox and KO BMDMs treated with or without LPS and IL-4 (n = 4–5), unpaired t test among different treatment groups. (C-F) Venn diagram and GSEA from RNA sequencing in Flox and KO BMDMs treated with or without LPS showed autophagy was one of the most important pathways. (G) Western blot of p62 (p = 0.014, Kruskal-Wallis test) and LC3 II/I (p = 0.001, Welch's ANOVA test) in flox, KO, and KO+3MA (5 mM) BMDMs treated with or without LPS (n = 4). (H) Western blot of IL-1β (p = 0.022, unpaired t test), IL-18 (p = 0.028, unpaired t test), TNF-α (p = 0.002, Unpaired t test with Welch's correction) and VEGFA (p = 0.045, unpaired t test) in KO and KO+3MA BMDMs treated with or without LPS (n = 3–4). (I) Levels of IL-1β (p = 0.049, unpaired t test with Welch's correction), IL-18 (p = 0.045, unpaired t test), TNF-α (p = 0.021, unpaired t test) and VEGFA (p < 0.001, unpaired t test) in supernatants of KO and KO+3MA BMDMs (n = 4–5). *p < 0.05, **p < 0.01, ***p < 0.001. “n value” refers to the number of biological replicates. ADAM8: A disintegrin and metallopeptidase domain 8. BMDMs: Bone marrow-derived macrophages. KO: Knockout. LPS: lipopolysaccharide. 3MA: 3-Methyladenine.

Journal: Journal of Advanced Research

Article Title: ADAM8 deficiency in macrophages promotes cardiac repair after myocardial infarction via ANXA2-mTOR-autophagy pathway

doi: 10.1016/j.jare.2024.07.037

Figure Lengend Snippet: ADAM8 regulates VEGFA secretion and inflammation in macrophages via autophagy pathway (A) Western blot of IL-1β, IL-18, TNF-α and VEGFA in Flox and KO BMDMs treated with or without LPS (1 μg/mL for 12 h) and IL-4 (20 ng/mL for 12 h) (n = 3–4), unpaired t test among different treatment groups. (B) Levels of IL-1β, IL-18, TNF-α and VEGFA in supernatants of Flox and KO BMDMs treated with or without LPS and IL-4 (n = 4–5), unpaired t test among different treatment groups. (C-F) Venn diagram and GSEA from RNA sequencing in Flox and KO BMDMs treated with or without LPS showed autophagy was one of the most important pathways. (G) Western blot of p62 (p = 0.014, Kruskal-Wallis test) and LC3 II/I (p = 0.001, Welch's ANOVA test) in flox, KO, and KO+3MA (5 mM) BMDMs treated with or without LPS (n = 4). (H) Western blot of IL-1β (p = 0.022, unpaired t test), IL-18 (p = 0.028, unpaired t test), TNF-α (p = 0.002, Unpaired t test with Welch's correction) and VEGFA (p = 0.045, unpaired t test) in KO and KO+3MA BMDMs treated with or without LPS (n = 3–4). (I) Levels of IL-1β (p = 0.049, unpaired t test with Welch's correction), IL-18 (p = 0.045, unpaired t test), TNF-α (p = 0.021, unpaired t test) and VEGFA (p < 0.001, unpaired t test) in supernatants of KO and KO+3MA BMDMs (n = 4–5). *p < 0.05, **p < 0.01, ***p < 0.001. “n value” refers to the number of biological replicates. ADAM8: A disintegrin and metallopeptidase domain 8. BMDMs: Bone marrow-derived macrophages. KO: Knockout. LPS: lipopolysaccharide. 3MA: 3-Methyladenine.

Article Snippet: Immunoprecipitation was also performed with the anti-ADAM8 antibody (IP: ADAM8) or IgG (IP: IgG) and RAW264.7 cells, and the protein lysates were used for LC-MS/MS (Applied Protein Technology Co. Ltd, Shanghai).

Techniques: Western Blot, RNA Sequencing, Derivative Assay, Knock-Out

ADAM8 binds to ANXA2 to govern function of macrophages (A) COIP/MS based on H293T presented by Venn diagram. (B) IP validation of ADAM8 binding to ANXA2 in RAW264.7 cells. (C) Representative colocalization immunofluorescence images of ADAM8 and ANXA2 in BMDMs treated with LPS (n = 3), Scale bar = 20 μm. (D) The expression of p-ANXA2 Ser26, ANXA2, and ADAM8 in IP groups with ANXA2 and ADAM8 based on flox and KO BMDMs (n = 3). (E-F) Western blot of p-ANXA2 Ser26, p-ANXA2 Tyr24, ANXA2, p-mTOR and mTOR in Flox and KO BMDMs treated with or without LPS (n = 3), One-way ANOVA and Tukey's test. (G) Western blot of p-ANXA2 Ser26/ANXA2 (p < 0.001), VEGFA (p = 0.008, Welch's ANOVA test), IL-1β (p = 0.002), IL-18 (p < 0.001), TNF-α (p = 0.038), p62 (p < 0.001), LC3II/I (p = 0.002), and p-mTOR/mTOR (p = 0.032) in NC, S26D and S26A BMDMs (n = 4–6), One-way ANOVA and Tukey's test. (H) Levels of IL-1β (p < 0.001), IL-18 (p = 0.005), TNF-α (p < 0.001) and VEGFA (p < 0.001) in supernatants of in NC, S26D and S26A BMDMs (n = 4), One-way ANOVA and Tukey's test. *p < 0.05, **p < 0.01, ***p < 0.001. “n value” refers to the number of biological replicates. ADAM8: A disintegrin and metallopeptidase domain 8. ANXA2: Annexin A2. BMDMs: Bone marrow-derived macrophages. KO: Knockout. LPS: lipopolysaccharide. mTOR: Mammalian target of rapamycin. S26D: Ser26 continuous activation. S26A: Ser26 continuous inhibition.

Journal: Journal of Advanced Research

Article Title: ADAM8 deficiency in macrophages promotes cardiac repair after myocardial infarction via ANXA2-mTOR-autophagy pathway

doi: 10.1016/j.jare.2024.07.037

Figure Lengend Snippet: ADAM8 binds to ANXA2 to govern function of macrophages (A) COIP/MS based on H293T presented by Venn diagram. (B) IP validation of ADAM8 binding to ANXA2 in RAW264.7 cells. (C) Representative colocalization immunofluorescence images of ADAM8 and ANXA2 in BMDMs treated with LPS (n = 3), Scale bar = 20 μm. (D) The expression of p-ANXA2 Ser26, ANXA2, and ADAM8 in IP groups with ANXA2 and ADAM8 based on flox and KO BMDMs (n = 3). (E-F) Western blot of p-ANXA2 Ser26, p-ANXA2 Tyr24, ANXA2, p-mTOR and mTOR in Flox and KO BMDMs treated with or without LPS (n = 3), One-way ANOVA and Tukey's test. (G) Western blot of p-ANXA2 Ser26/ANXA2 (p < 0.001), VEGFA (p = 0.008, Welch's ANOVA test), IL-1β (p = 0.002), IL-18 (p < 0.001), TNF-α (p = 0.038), p62 (p < 0.001), LC3II/I (p = 0.002), and p-mTOR/mTOR (p = 0.032) in NC, S26D and S26A BMDMs (n = 4–6), One-way ANOVA and Tukey's test. (H) Levels of IL-1β (p < 0.001), IL-18 (p = 0.005), TNF-α (p < 0.001) and VEGFA (p < 0.001) in supernatants of in NC, S26D and S26A BMDMs (n = 4), One-way ANOVA and Tukey's test. *p < 0.05, **p < 0.01, ***p < 0.001. “n value” refers to the number of biological replicates. ADAM8: A disintegrin and metallopeptidase domain 8. ANXA2: Annexin A2. BMDMs: Bone marrow-derived macrophages. KO: Knockout. LPS: lipopolysaccharide. mTOR: Mammalian target of rapamycin. S26D: Ser26 continuous activation. S26A: Ser26 continuous inhibition.

Article Snippet: Immunoprecipitation was also performed with the anti-ADAM8 antibody (IP: ADAM8) or IgG (IP: IgG) and RAW264.7 cells, and the protein lysates were used for LC-MS/MS (Applied Protein Technology Co. Ltd, Shanghai).

Techniques: Biomarker Discovery, Binding Assay, Immunofluorescence, Expressing, Western Blot, Derivative Assay, Knock-Out, Activation Assay, Inhibition

Prognostic and immune cell correlation analysis of PRDEGs. a – j KM curves were independently analyzed at the transcriptome level to determine the prognostic value of these PRDEGs ( a – j : HEYL, FSTL3, HOMER3, CALB2, FABP4, MEGF6, ADAM8, TIMP1, CLCA1, NOS2). k The results of the correlation analysis between the PRDEGs and each infiltrating immunocyte are displayed in a heatmap

Journal: Discover Oncology

Article Title: Disulfidptosis-based molecular clustering and prognostic signatures predict patient survival and the immune landscape in patients with colon cancer

doi: 10.1007/s12672-025-02142-w

Figure Lengend Snippet: Prognostic and immune cell correlation analysis of PRDEGs. a – j KM curves were independently analyzed at the transcriptome level to determine the prognostic value of these PRDEGs ( a – j : HEYL, FSTL3, HOMER3, CALB2, FABP4, MEGF6, ADAM8, TIMP1, CLCA1, NOS2). k The results of the correlation analysis between the PRDEGs and each infiltrating immunocyte are displayed in a heatmap

Article Snippet: Primary antibody information for the PRDEGs was as follows: HEYL (GB114972, Servicebio, 1:1000), NOS2 (GB11119, Servicebio, 1:500), FABP4 (GB115466, Servicebio, 1:200), TIMP1 (GB14158, Servicebio, 1:100), MEGF6 (bs-18776R, Bioss, 1:200), FSTL3 (A10279, abclonal, 1:200), CALB2 (66496-1-Ig, Proteintech, 1:4000), HOMER3 (16624-1-AP, Proteintech, 1:200), ADAM8 (23778-1-AP, Proteintech, 1:50), and CLCA1 (A15041, abclonal, 1:200).

Techniques:

Immunohistochemical analysis of PRDEGs based on the SI. a Expression of PRDEGs at the protein level. ( b – d ) KM curves showing that patients with low expression of ADAM8 ( d ) and FSTL3 ( c ) and high expression of FABP4 ( b ) had a better prognosis

Journal: Discover Oncology

Article Title: Disulfidptosis-based molecular clustering and prognostic signatures predict patient survival and the immune landscape in patients with colon cancer

doi: 10.1007/s12672-025-02142-w

Figure Lengend Snippet: Immunohistochemical analysis of PRDEGs based on the SI. a Expression of PRDEGs at the protein level. ( b – d ) KM curves showing that patients with low expression of ADAM8 ( d ) and FSTL3 ( c ) and high expression of FABP4 ( b ) had a better prognosis

Article Snippet: Primary antibody information for the PRDEGs was as follows: HEYL (GB114972, Servicebio, 1:1000), NOS2 (GB11119, Servicebio, 1:500), FABP4 (GB115466, Servicebio, 1:200), TIMP1 (GB14158, Servicebio, 1:100), MEGF6 (bs-18776R, Bioss, 1:200), FSTL3 (A10279, abclonal, 1:200), CALB2 (66496-1-Ig, Proteintech, 1:4000), HOMER3 (16624-1-AP, Proteintech, 1:200), ADAM8 (23778-1-AP, Proteintech, 1:50), and CLCA1 (A15041, abclonal, 1:200).

Techniques: Immunohistochemical staining, Expressing

Immunohistochemical images (ADAM8, CLCA1, FABP4, FSTL3, HOMER3)

Journal: Discover Oncology

Article Title: Disulfidptosis-based molecular clustering and prognostic signatures predict patient survival and the immune landscape in patients with colon cancer

doi: 10.1007/s12672-025-02142-w

Figure Lengend Snippet: Immunohistochemical images (ADAM8, CLCA1, FABP4, FSTL3, HOMER3)

Article Snippet: Primary antibody information for the PRDEGs was as follows: HEYL (GB114972, Servicebio, 1:1000), NOS2 (GB11119, Servicebio, 1:500), FABP4 (GB115466, Servicebio, 1:200), TIMP1 (GB14158, Servicebio, 1:100), MEGF6 (bs-18776R, Bioss, 1:200), FSTL3 (A10279, abclonal, 1:200), CALB2 (66496-1-Ig, Proteintech, 1:4000), HOMER3 (16624-1-AP, Proteintech, 1:200), ADAM8 (23778-1-AP, Proteintech, 1:50), and CLCA1 (A15041, abclonal, 1:200).

Techniques: Immunohistochemical staining

Univariate Cox regression analysis of expression of PRDEGs at the protein level and clinical parameters

Journal: Discover Oncology

Article Title: Disulfidptosis-based molecular clustering and prognostic signatures predict patient survival and the immune landscape in patients with colon cancer

doi: 10.1007/s12672-025-02142-w

Figure Lengend Snippet: Univariate Cox regression analysis of expression of PRDEGs at the protein level and clinical parameters

Article Snippet: Primary antibody information for the PRDEGs was as follows: HEYL (GB114972, Servicebio, 1:1000), NOS2 (GB11119, Servicebio, 1:500), FABP4 (GB115466, Servicebio, 1:200), TIMP1 (GB14158, Servicebio, 1:100), MEGF6 (bs-18776R, Bioss, 1:200), FSTL3 (A10279, abclonal, 1:200), CALB2 (66496-1-Ig, Proteintech, 1:4000), HOMER3 (16624-1-AP, Proteintech, 1:200), ADAM8 (23778-1-AP, Proteintech, 1:50), and CLCA1 (A15041, abclonal, 1:200).

Techniques: Expressing

Prognostic and immune cell correlation analysis of PRDEGs. a – j KM curves were independently analyzed at the transcriptome level to determine the prognostic value of these PRDEGs ( a – j : HEYL, FSTL3, HOMER3, CALB2, FABP4, MEGF6, ADAM8, TIMP1, CLCA1, NOS2). k The results of the correlation analysis between the PRDEGs and each infiltrating immunocyte are displayed in a heatmap

Journal: Discover Oncology

Article Title: Disulfidptosis-based molecular clustering and prognostic signatures predict patient survival and the immune landscape in patients with colon cancer

doi: 10.1007/s12672-025-02142-w

Figure Lengend Snippet: Prognostic and immune cell correlation analysis of PRDEGs. a – j KM curves were independently analyzed at the transcriptome level to determine the prognostic value of these PRDEGs ( a – j : HEYL, FSTL3, HOMER3, CALB2, FABP4, MEGF6, ADAM8, TIMP1, CLCA1, NOS2). k The results of the correlation analysis between the PRDEGs and each infiltrating immunocyte are displayed in a heatmap

Article Snippet: Primary antibody information for the PRDEGs was as follows: HEYL (GB114972, Servicebio, 1:1000), NOS2 (GB11119, Servicebio, 1:500), FABP4 (GB115466, Servicebio, 1:200), TIMP1 (GB14158, Servicebio, 1:100), MEGF6 (bs-18776R, Bioss, 1:200), FSTL3 (A10279, abclonal, 1:200), CALB2 (66496-1-Ig, Proteintech, 1:4000), HOMER3 (16624-1-AP, Proteintech, 1:200), ADAM8 (23778-1-AP, Proteintech, 1:50), and CLCA1 (A15041, abclonal, 1:200).

Techniques:

Immunohistochemical images (ADAM8, CLCA1, FABP4, FSTL3, HOMER3)

Journal: Discover Oncology

Article Title: Disulfidptosis-based molecular clustering and prognostic signatures predict patient survival and the immune landscape in patients with colon cancer

doi: 10.1007/s12672-025-02142-w

Figure Lengend Snippet: Immunohistochemical images (ADAM8, CLCA1, FABP4, FSTL3, HOMER3)

Article Snippet: Primary antibody information for the PRDEGs was as follows: HEYL (GB114972, Servicebio, 1:1000), NOS2 (GB11119, Servicebio, 1:500), FABP4 (GB115466, Servicebio, 1:200), TIMP1 (GB14158, Servicebio, 1:100), MEGF6 (bs-18776R, Bioss, 1:200), FSTL3 (A10279, abclonal, 1:200), CALB2 (66496-1-Ig, Proteintech, 1:4000), HOMER3 (16624-1-AP, Proteintech, 1:200), ADAM8 (23778-1-AP, Proteintech, 1:50), and CLCA1 (A15041, abclonal, 1:200).

Techniques: Immunohistochemical staining

Univariate Cox regression analysis of expression of PRDEGs at the protein level and clinical parameters

Journal: Discover Oncology

Article Title: Disulfidptosis-based molecular clustering and prognostic signatures predict patient survival and the immune landscape in patients with colon cancer

doi: 10.1007/s12672-025-02142-w

Figure Lengend Snippet: Univariate Cox regression analysis of expression of PRDEGs at the protein level and clinical parameters

Article Snippet: Primary antibody information for the PRDEGs was as follows: HEYL (GB114972, Servicebio, 1:1000), NOS2 (GB11119, Servicebio, 1:500), FABP4 (GB115466, Servicebio, 1:200), TIMP1 (GB14158, Servicebio, 1:100), MEGF6 (bs-18776R, Bioss, 1:200), FSTL3 (A10279, abclonal, 1:200), CALB2 (66496-1-Ig, Proteintech, 1:4000), HOMER3 (16624-1-AP, Proteintech, 1:200), ADAM8 (23778-1-AP, Proteintech, 1:50), and CLCA1 (A15041, abclonal, 1:200).

Techniques: Expressing