Journal: Journal of Biochemical and Molecular Toxicology

Article Title: Tumor‐Associated Mast Cells Drive Immunosuppression and Resistance to Anti‐PD‐1 Therapy in Hepatocellular Carcinoma Through CCL17/CCR4‐mediated Regulatory T‐Cell Recruitment

doi: 10.1002/jbt.71127

Figure Lengend Snippet: HCC cells promote MC migration and sustain MC proliferation. (A, B) Timer3.0 database analysis of SCF expression in tumors and correlation analysis between SCF and MCs; (C) ELISA detection of SCF levels in tumor tissues and adjacent tissues; (D) Correlation analysis between TAMC density and SCF concentration in HCC tumor tissues; (E): Flow cytometry identification of expanded hMCs at week 4; (F, G) Transwell assay for hMC migration capacity; (H, I) Flow cytometry for hMC proliferation; (J, K) Flow cytometry for hMC cell cycle. Data are presented as mean ± SD. Figure (B) used independent samples t ‐tests ( n = 59). (D) Spearman 's rank correlation analysis ( n = 59); (G/I/K), one‐way ANOVA with Tukey's post‐hoc multiple comparison test ( N = 3). * indicates p < 0.05; *** indicates p < 0.001.

Article Snippet: The following kits were utilized: human SCF ELISA Kit (E‐EL‐H1299, Elabscience), human CCL1 ELISA kit (ab314600, Abcam), human CCL5 ELISA kit (ab174446, Abcam), human CCL17 ELISA kit (ab183366, Abcam), Human CCL22 ELISA Kit (ab242931, Abcam), Human CCL28 ELISA Kit (ab99988, Abcam), Human Granzyme B ELISA Kit (E‐EL‐H1617, Elabscience), Human Interferon‐γ (IFN‐γ) ELISA Kit (E‐EL‐H0108, Elabscience), Human TNF‐α ELISA Kit (E‐EL‐H0109, Elabscience), Human Perforin 1 ELISA Kit (E‐EL‐H1123, Elabscience), Mouse Granzyme B ELISA Kit (E‐EL‐M0594, Elabscience), Mouse IFN‐γ ELISA Kit (E‐EL‐M0048, Elabscience), Mouse TNF‐α ELISA Kit (E‐EL‐M3063, Elabscience), and Mouse Perforin ELISA Kit (E‐EL‐M0890, Elabscience).

Techniques: Migration, Analysis, Expressing, Enzyme-linked Immunosorbent Assay, Concentration Assay, Flow Cytometry, Transwell Assay, Comparison, X-Ray Photoelectron Spectroscopy

Journal: Journal of Biochemical and Molecular Toxicology

Article Title: Tumor‐Associated Mast Cells Drive Immunosuppression and Resistance to Anti‐PD‐1 Therapy in Hepatocellular Carcinoma Through CCL17/CCR4‐mediated Regulatory T‐Cell Recruitment

doi: 10.1002/jbt.71127

Figure Lengend Snippet: TAMCs recruit Tregs into the tumor microenvironment by secreting chemokines. (A) ELISA detection of CCL1, CCL5, CCL17, CCL22, and CCL28 secretion by TAMCs (after subtracting CCL1, CCL5, CCL17, CCL22, and CCL28 levels in TCCS); (B) ELISA detection of CCL17 levels in tumor tissues and adjacent tissues; (C) Correlation analysis between TAMCs and CCL17 concentration; D: Correlation analysis between Tregs and CCL17 concentration; (E, F) Transwell migration assay to assess Treg migration capacity. Data are presented as mean ± SD. (A, B), independent samples t ‐tests ( N = 3); (C, D) Spearman 's rank correlation analysis ( n = 59); (F), one‐way ANOVA and Tukey's multiple comparison test ( N = 3). * indicates p < 0.05, ** indicates p < 0.01, *** indicates p < 0.001.

Article Snippet: The following kits were utilized: human SCF ELISA Kit (E‐EL‐H1299, Elabscience), human CCL1 ELISA kit (ab314600, Abcam), human CCL5 ELISA kit (ab174446, Abcam), human CCL17 ELISA kit (ab183366, Abcam), Human CCL22 ELISA Kit (ab242931, Abcam), Human CCL28 ELISA Kit (ab99988, Abcam), Human Granzyme B ELISA Kit (E‐EL‐H1617, Elabscience), Human Interferon‐γ (IFN‐γ) ELISA Kit (E‐EL‐H0108, Elabscience), Human TNF‐α ELISA Kit (E‐EL‐H0109, Elabscience), Human Perforin 1 ELISA Kit (E‐EL‐H1123, Elabscience), Mouse Granzyme B ELISA Kit (E‐EL‐M0594, Elabscience), Mouse IFN‐γ ELISA Kit (E‐EL‐M0048, Elabscience), Mouse TNF‐α ELISA Kit (E‐EL‐M3063, Elabscience), and Mouse Perforin ELISA Kit (E‐EL‐M0890, Elabscience).

Techniques: Enzyme-linked Immunosorbent Assay, Analysis, Concentration Assay, Transwell Migration Assay, Migration, Comparison

Journal: Journal of Biochemical and Molecular Toxicology

Article Title: Tumor‐Associated Mast Cells Drive Immunosuppression and Resistance to Anti‐PD‐1 Therapy in Hepatocellular Carcinoma Through CCL17/CCR4‐mediated Regulatory T‐Cell Recruitment

doi: 10.1002/jbt.71127

Figure Lengend Snippet: The TAMCs‐Tregs axis inhibits CD8 + T cell function. (A, B) Flow cytometry analysis of Treg‐mediated suppression of CD8 + T cell proliferation; (C–F) ELISA for Granzyme B, IFN‐γ, TNF‐α, and Perforin in supernatants from CD8 + T cell proliferation assays. Data are presented as mean ± SD. One‐way ANOVA was performed, followed by Tukey's multiple comparison test for post‐hoc analysis ( N = 3). * indicates p < 0.05, ** indicates p < 0.01, *** indicates p < 0.001.

Article Snippet: The following kits were utilized: human SCF ELISA Kit (E‐EL‐H1299, Elabscience), human CCL1 ELISA kit (ab314600, Abcam), human CCL5 ELISA kit (ab174446, Abcam), human CCL17 ELISA kit (ab183366, Abcam), Human CCL22 ELISA Kit (ab242931, Abcam), Human CCL28 ELISA Kit (ab99988, Abcam), Human Granzyme B ELISA Kit (E‐EL‐H1617, Elabscience), Human Interferon‐γ (IFN‐γ) ELISA Kit (E‐EL‐H0108, Elabscience), Human TNF‐α ELISA Kit (E‐EL‐H0109, Elabscience), Human Perforin 1 ELISA Kit (E‐EL‐H1123, Elabscience), Mouse Granzyme B ELISA Kit (E‐EL‐M0594, Elabscience), Mouse IFN‐γ ELISA Kit (E‐EL‐M0048, Elabscience), Mouse TNF‐α ELISA Kit (E‐EL‐M3063, Elabscience), and Mouse Perforin ELISA Kit (E‐EL‐M0890, Elabscience).

Techniques: Cell Function Assay, Flow Cytometry, Analysis, Enzyme-linked Immunosorbent Assay, Comparison

Journal: Journal of Biochemical and Molecular Toxicology

Article Title: Tumor‐Associated Mast Cells Drive Immunosuppression and Resistance to Anti‐PD‐1 Therapy in Hepatocellular Carcinoma Through CCL17/CCR4‐mediated Regulatory T‐Cell Recruitment

doi: 10.1002/jbt.71127

Figure Lengend Snippet: Combination therapy of anti‐PD‐1 antibody and CCR4 antagonists reshapes the tumor immune microenvironment in HCC. (A–F) Flow cytometry analysis of immune cell proportions (MCs, Tregs, CD8 + T cells) in tumor tissue; G–K: ELISA detection of Granzyme B, IFN‐γ, TNF‐α, Perforin, and CCL17 levels. Data are presented as mean ± SD. One‐way ANOVA was performed, followed by Tukey's multiple comparison test for post‐hoc analysis ( n = 6). * indicates p < 0.05, ** indicates p < 0.01, *** indicates p < 0.001.

Article Snippet: The following kits were utilized: human SCF ELISA Kit (E‐EL‐H1299, Elabscience), human CCL1 ELISA kit (ab314600, Abcam), human CCL5 ELISA kit (ab174446, Abcam), human CCL17 ELISA kit (ab183366, Abcam), Human CCL22 ELISA Kit (ab242931, Abcam), Human CCL28 ELISA Kit (ab99988, Abcam), Human Granzyme B ELISA Kit (E‐EL‐H1617, Elabscience), Human Interferon‐γ (IFN‐γ) ELISA Kit (E‐EL‐H0108, Elabscience), Human TNF‐α ELISA Kit (E‐EL‐H0109, Elabscience), Human Perforin 1 ELISA Kit (E‐EL‐H1123, Elabscience), Mouse Granzyme B ELISA Kit (E‐EL‐M0594, Elabscience), Mouse IFN‐γ ELISA Kit (E‐EL‐M0048, Elabscience), Mouse TNF‐α ELISA Kit (E‐EL‐M3063, Elabscience), and Mouse Perforin ELISA Kit (E‐EL‐M0890, Elabscience).

Techniques: Flow Cytometry, Analysis, Immune Cell, Tissue, Enzyme-linked Immunosorbent Assay, Comparison

Journal: Journal of Biochemical and Molecular Toxicology

Article Title: Tumor‐Associated Mast Cells Drive Immunosuppression and Resistance to Anti‐PD‐1 Therapy in Hepatocellular Carcinoma Through CCL17/CCR4‐mediated Regulatory T‐Cell Recruitment

doi: 10.1002/jbt.71127

Figure Lengend Snippet: Validation of the synergistic antitumor efficacy of the triple‐drug therapy in an in situ Hepa1‐6 HCC model. (A) Representative images of in situ tumors; (B): Tumor volume in each group; (C) Tumor weight in each group; (D) Average ratio of liver weight to body weight; (E, F): Ki67 immunohistochemical staining and quantitative analysis; (G–L) Flow cytometry detection of the proportions of various immune cells (mast cells, Tregs, CD8 + T cells); (M) ELISA detection of Granzyme B, IFN‐γ, TNF‐α, and perforin levels. Data are presented as mean ± SD. One‐way ANOVA was performed, followed by Tukey's multiple comparison test for post‐hoc analysis ( n = 6).

Article Snippet: The following kits were utilized: human SCF ELISA Kit (E‐EL‐H1299, Elabscience), human CCL1 ELISA kit (ab314600, Abcam), human CCL5 ELISA kit (ab174446, Abcam), human CCL17 ELISA kit (ab183366, Abcam), Human CCL22 ELISA Kit (ab242931, Abcam), Human CCL28 ELISA Kit (ab99988, Abcam), Human Granzyme B ELISA Kit (E‐EL‐H1617, Elabscience), Human Interferon‐γ (IFN‐γ) ELISA Kit (E‐EL‐H0108, Elabscience), Human TNF‐α ELISA Kit (E‐EL‐H0109, Elabscience), Human Perforin 1 ELISA Kit (E‐EL‐H1123, Elabscience), Mouse Granzyme B ELISA Kit (E‐EL‐M0594, Elabscience), Mouse IFN‐γ ELISA Kit (E‐EL‐M0048, Elabscience), Mouse TNF‐α ELISA Kit (E‐EL‐M3063, Elabscience), and Mouse Perforin ELISA Kit (E‐EL‐M0890, Elabscience).

Techniques: Biomarker Discovery, In Situ, Immunohistochemical staining, Staining, Analysis, Flow Cytometry, Enzyme-linked Immunosorbent Assay, Comparison