anti cd8 t cell depletion antibody  (Bio X Cell)

Journal: Redox biology

Article Title: Radiation-induced upregulation of itaconate in macrophages promotes the radioresistance of non-small cell lung cancer by stabilizing NRF2 protein and suppressing immune response.

doi: 10.1016/j.redox.2025.103711

Figure Lengend Snippet: Fig. 5. Knockout of Acod1 in macrophages further enhance the activation of tumor immune microenvironment by radiotherapy (A) Gating strategy for detection of NK, CD4+ and CD8+ T cells by flow cytometry. (B–G) Flow cytometry analysis of CD45+ immune cells (B), CD3+ (C), CD4+ (D), CD8+ (E) T cells, the ratio of CD4+/CD8+ T cells (F), and NK cells (G) in the TME of LLC subcutaneous tumor model in mice treated with radiation (8 Gy x 3) in Acod1f/f Lyz2cre− (HO−) and Acod1f/f Lyz2cre+ (HO+) mice (n = 6). (H) Representative immunofluorescence staining of CD8 (green) in the TME at the end of experiment in Acod1f/f Lyz2cre−(HO−) and Acod1f/f Lyz2cre+ (HO+) mice. (I–J) Flow cytometry analysis of CD107a+ (I), and IFN-γ+ CD8+ T cells (J) in the TME of LLC subcutaneous tumor model described above (n = 6). *p < 0.05; **p < 0.01; ***p < 0.001; ns, not statistically significant.

Article Snippet: CD8+ T cell depletion Anti-mouse CD8 monoclonal antibody (clone 2.43) (#A2102, Selleck, China) was intraperitoneally injected at a dose of 200 μg per mouse, and 150 μg per mouse was injected 3 days later.

Techniques: Knock-Out, Activation Assay, Flow Cytometry, Immunofluorescence, Staining

Journal: Redox biology

Article Title: Radiation-induced upregulation of itaconate in macrophages promotes the radioresistance of non-small cell lung cancer by stabilizing NRF2 protein and suppressing immune response.

doi: 10.1016/j.redox.2025.103711

Figure Lengend Snippet: Fig. 6. The antitumor effect of Acod1 knockout in macrophages combined with radiotherapy partially depends on CD8+ T cells (A) CD8+ T cells clearance efficiency in mouse spleen detected by flow cytometry (n = 3). (B) CD8+ T cells clearance efficiency in mouse peripheral blood detected by flow cytometry (n = 3). (C) Tumor growth curves of LLC subcutaneous tumor in Acod1f/f Lyz2cre−(HO−) and Acod1f/f Lyz2cre+ (HO+) mice treated with 8 Gy x 3 radiotherapy and IgG or neutralizing CD8 antibody (α-CD8) (n = 6). (D–E) Tumor weight (D) and tumor images (E) on day 16 after treatment in different groups (n = 6). (F) The schematic diagram depicting that macrophages up-regulate the expression of Acod1 through activating NF-κB pathway after radiotherapy, thereby regulating the radiosensitivity of tumor cells and immune cell infiltration. *p < 0.05; **p < 0.01; ***p < 0.001; ns, not statistically significant.

Article Snippet: CD8+ T cell depletion Anti-mouse CD8 monoclonal antibody (clone 2.43) (#A2102, Selleck, China) was intraperitoneally injected at a dose of 200 μg per mouse, and 150 μg per mouse was injected 3 days later.

Techniques: Knock-Out, Flow Cytometry, Expressing

Journal: Journal for immunotherapy of cancer

Article Title: Differential requirements for CD4+ T cells in the efficacy of the anti-PD-1+LAG-3 and anti-PD-1+CTLA-4 combinations in melanoma flank and brain metastasis models.

doi: 10.1136/jitc-2023-007239

Figure Lengend Snippet: Figure 2 The anti-PD-1+LAG-3 and the anti-PD-1+CTLA-4 combinations suppress melanoma brain metastases in the SM1 mouse melanoma model. (A) The ICI doublets were more effective than single agent ICI in the brain tumors of SM1 mouse melanoma model with 2/10 mice relapsing in each combination group. Mice received 200 µg/100 µL i.p. doses of single agents and doublets every 5 days for 80 days. (B) MRI T2 sequence of brain of mice with SM1 MBM tumors demonstrating a lack of tumor regrowth after holding the therapy for 28 days. MRIs were generated using standard T2-weighted Turbo Spring Echo. Brain tumors were manually contoured on all MRI slices using ImageJ. A custom program was used to extract voxels within the manually drawn contours and compute total volume burden, in mm3. (C) Monitoring melanoma brain metastases by MRI. Time course of MRI T2 sequences of brain of mouse melanoma brain metastases following treatment with IgG, anti-PD- 1+LAG-3 or anti-PD-1+CTLA-4. MRIs were taken twice a week and were generated using standard T2-weighted Turbo Spring Echo. Brain tumors were manually contoured on all MRI slices using ImageJ. A custom MATLAB program was used to extract voxels within the manually drawn contours and compute total volume burden, in mm3. (D–F) Kinetics of CD4+ and CD8+ T cell, Myeloid-derived suppressor cells (MDSCs) and macrophage infiltrate following treatment with each ICI doublet over a period of 28 days in SM1 brain tumors. Tumors were collected at days 7 and 14 for IgG and at days 7, 14, and 28 after treatment initiation for the doublets. The results were represented as average ±SEM with 10 mice per group for all the growth curve experiments. The results were represented as average±SEM of 3 mice per group for panels D–F. Statistical significance was assessed with one-way ANOVA test (*p<0.05, **p<0.01, ***p<0.001). ANOVA, analysis of variance; ICI, immune checkpoint inhibitor.

Article Snippet: CD4+ and CD8+ T cell depletion CD4- specific antibody (clone YTS191; cat. #BE0003- 1; Bio X Cell) and CD8a- specific antibody (clone YTS169.4; cat. #BE0117; Bio X Cell) from Bio X Cell were used to deplete CD4+ T cells and CD8+ T cells, respectively.

Techniques: Sequencing, Generated, Derivative Assay

Journal: Journal for immunotherapy of cancer

Article Title: Differential requirements for CD4+ T cells in the efficacy of the anti-PD-1+LAG-3 and anti-PD-1+CTLA-4 combinations in melanoma flank and brain metastasis models.

doi: 10.1136/jitc-2023-007239

Figure Lengend Snippet: Figure 3 Immune landscape of flank and brain tumors in SM1 mouse model following the treatment with single agent and combination ICI therapy. A) t-SNE plots showing major cell types identified in relapsed brain tumors and responding flank SM1 tumors following treatment with single agent and combination ICI. (B) Proportion of each cell type in the responding flank tumors from the indicted treatment groups. (C) Proportion of different T-cell clusters identified in the responding flank tumors from the indicated treatment groups. (D) Heatmap showing expression of activation/exhaustion markers across the identified T-cell clusters. (E) Violin plots showing expression of T cell activation markers and immune checkpoints in each T-cell cluster. (F) IHC analysis of tumors treated with anti-PD-1+LAG-3 or anti-PD-1+CTLA-4 demonstrates increased CD4+and CD8+ T cell infiltration in responding flank tumors. The samples were stained with anti-CD4 and anti-CD8 by IHC. Tumors were harvested at day 9 for IgG and single agents and at day 19 for anti-PD-1+LAG-3 and day 25 for anti-PD-1+CTLA-4 following the initiation of treatment. ICI, immune checkpoint inhibitor.

Article Snippet: CD4+ and CD8+ T cell depletion CD4- specific antibody (clone YTS191; cat. #BE0003- 1; Bio X Cell) and CD8a- specific antibody (clone YTS169.4; cat. #BE0117; Bio X Cell) from Bio X Cell were used to deplete CD4+ T cells and CD8+ T cells, respectively.

Techniques: Expressing, Activation Assay, Staining

Journal: Journal for immunotherapy of cancer

Article Title: Differential requirements for CD4+ T cells in the efficacy of the anti-PD-1+LAG-3 and anti-PD-1+CTLA-4 combinations in melanoma flank and brain metastasis models.

doi: 10.1136/jitc-2023-007239

Figure Lengend Snippet: Figure 4 Anti-PD-1+LAG-3 and anti-PD-1+CTLA-4 differentially polarize CD4+T cells in the SM1 mouse melanoma model. (A) Proportion of T cell clusters and three major subsets of CD4+T cells identified in the responding flank tumors from the indicated treatment groups. (B) Expression of immune checkpoints and markers for T helper subsets and T regulatory cells in the three major subsets of CD4+T cells identified by violin plots. (C) Percentage of CD4+T cells and activated CD69+CD4+ T cells in the responding flank tumors treated with combination of anti-PD-1+LAG-3 and anti-PD-1+CTLA-4 identified by flow cytometry. Tumors were harvested at the endpoint following the treatment initiation. (D) Flow cytometry histogram plot showing activated CD4+T cells in tumors treated with IgG, anti-PD-1+LAG-3 or anti-PD-1+CTLA-4 as evidenced by cell surface CD69 and CD4 staining. (E) Treatment with anti-PD-1+CTLA-4 increased Tregs infiltration in the responding flank tumors and relapsing brain tumors. The samples were stained with anti-FOXP3 by IHC. Tumors were harvested at day 9 for IgG and single agents and at day 19 for anti-PD-1+LAG-3 and day 25 for anti-PD-1+CTLA-4 following the initiation of treatment for sc-RNAseq and IHC. The results were represented as average±SEM of 3 mice per group for panels C and D. Statistical significance was assessed with one-way ANOVA test (***p<0.001). ANOVA, analysis of variance ; IHC, immunohistochemistry.

Article Snippet: CD4+ and CD8+ T cell depletion CD4- specific antibody (clone YTS191; cat. #BE0003- 1; Bio X Cell) and CD8a- specific antibody (clone YTS169.4; cat. #BE0117; Bio X Cell) from Bio X Cell were used to deplete CD4+ T cells and CD8+ T cells, respectively.

Techniques: Expressing, Flow Cytometry, Staining, Immunohistochemistry

Journal: Journal for immunotherapy of cancer

Article Title: Differential requirements for CD4+ T cells in the efficacy of the anti-PD-1+LAG-3 and anti-PD-1+CTLA-4 combinations in melanoma flank and brain metastasis models.

doi: 10.1136/jitc-2023-007239

Figure Lengend Snippet: Figure 5 Antitumor responses to anti-PD-1+LAG-3 are dependent on CD4+T helper function. (A, B) Depletion of CD4+T cells demonstrates that responses to anti-PD-1+LAG-3 but not anti-PD-1+CTLA-4 are dependent on CD4+T cells in both SM1 flank and brain tumor models. The results were represented as an average±SEM of 5 mice per group. (C) Decreased percentage of tumor-infiltrating CD69+CD8+ T cells and CD8+T cells in SM1 flank tumors, respectively, following CD4+T cell depletion and anti-PD-1+LAG-3 treatment. Increased percentages of CD69+CD8+ T cells and total CD8+T cells were seen in SM1 flank tumors following CD4+T cell depletion and anti-PD-1+CTLA-4 treatment. Tumors were collected at day 10 after the treatment initiation. (D) Antitumor responses of the PD-1+LAG-3 but not the PD-1+CTLA-4 combination are dependent on CD4+T cells in B16 flank tumors. Results show the average±SEM of 5 mice per group. (E) Decreased percentage of tumor-infiltrating CD69+CD8+ T cells and CD8+T cells in B flank tumors, respectively, following CD4+T cell depletion and anti-PD-1+LAG-3 treatment. Increased percentages of CD69+CD8+ T cells and total CD8+T cells were seen in B16 flank tumors following CD4+T cell depletion and anti-PD-1+CTLA-4 treatment. Tumors were harvested for flow cytometry analysis at day 5 following the start of the treatment. (F) Responses to Anti-PD-1+LAG-3 but not Anti-PD-1+CTLA-4 are dependent on CD4+T cells in the flank tumors of D4M- UV2 mouse melanoma model. The results were represented as an average±SEM of 5 mice per group. (G) G) Percentage of CD69+CD8+ and CD8+ T cells decrease in the D4M-UV2 flank tumors significantly, following CD4+T cell depletion and Anti-PD- 1+LAG-3 treatment. Tumors were collected for flow cytometry analysis at day 16 of treatment for IgG and day 19 of treatment for the doublets. (H, I) ELISPOT assays showing decreased levels of IFNγ production following CD4+T cell depletion and treatment with the PD-1+LAG-3 combination in both the SM1 and B16 mouse melanoma models. The results were represented as average±SEM of 3 mice per group for panels D-G. Statistical significance was assessed with one-way ANOVA test (*p<0.05, **p<0.01, ***p<0.001). ANOVA, analysis of variance.

Article Snippet: CD4+ and CD8+ T cell depletion CD4- specific antibody (clone YTS191; cat. #BE0003- 1; Bio X Cell) and CD8a- specific antibody (clone YTS169.4; cat. #BE0117; Bio X Cell) from Bio X Cell were used to deplete CD4+ T cells and CD8+ T cells, respectively.

Techniques: Flow Cytometry, Enzyme-linked Immunospot