Journal: Cell Death & Disease

Article Title: Targeting tumor intrinsic TAK1 engages TNF-α-driven cell death through distinct mechanisms and enhances cancer immunotherapy

doi: 10.1038/s41419-025-08013-0

Figure Lengend Snippet: A MC38 cells lacking Tak1 or B2m expression were pulsed with the MHC-I restricted chicken ovalbumin SIINFEKL peptide and subsequently cultured with increasing ratios of OT-I CD8 T cells, previously activated for 48 h by CD3 and CD28 stimulation. Tumor cell viability was assessed 24 h later by flow cytometry, using CD8 surface staining to discriminate T cells from tumor cells. B – D CD8 T cells isolated from naïve mouse spleens (Balb/c for CT-26 and EMT6, C57BL/6 for MC38) were activated by CD3 and CD28 stimulation in vitro and subsequently cultured with ( B ) MC38, ( C ) CT-26, or ( D ) EMT6 tumor cells lacking Tak1 expression. Viability was assessed using DRAQ7 uptake, imaging cells every 2 h for the indicated times. E – G Tak1- deficient MC38 ( E ) or CT-26 ( F ) cells were cultured in media derived from 48 h activated CD-8 T cells. Viability was assessed via DRAQ7 uptake. G Conditioned media from 48 h activated CD8 T cells was added to EMT6 cells deficient for Tak1 , Tnfr1 , or Tak1 and Tnfr1 and viability was assessed via DRAQ7 uptake. All panels represent the mean +/− SD from a single experiment, n = 2 independent experiments.

Article Snippet: For the CD8 depletion protocol, mice were injected with anti-mouse CD8 (BioXCell; BP0061, New Haven, CT) or IgG2a Isotype control (BioXCell; BP 00085) at 10 mg/kg i.p. at the dose schedule of Day -2, Day -1 prior to flank inoculation of cells, then again on Days 4, 8, and 11 post inoculation.

Techniques: Expressing, Cell Culture, Flow Cytometry, Staining, Isolation, In Vitro, Imaging, Derivative Assay

Journal: Cell Death & Disease

Article Title: Targeting tumor intrinsic TAK1 engages TNF-α-driven cell death through distinct mechanisms and enhances cancer immunotherapy

doi: 10.1038/s41419-025-08013-0

Figure Lengend Snippet: A Immunocompetent mice (C57BL/6, n = 15/group) were implanted with pooled clones from Tak1 -deficient or parental MC38 cells and tumor growth was monitored. B , C Immunocompetent mice (BALB/c, n = 8/group) were implanted with Tak1 -deficient CT-26 cells ( B ) and treated at the indicated days with an α-PD-1 antibody ( C , 10 mg/kg) and tumor growth was monitored. D Immunodeficient mice (NSG, n = 10/group) were implanted with Tak1 -deficient CT-26 cells and tumor growth was monitored. E Quantification of the percentage of complete tumor clearance from panels ( B – D ). F Balb/c mice implanted with Tak1 -deficient CT-26 tumors and exhibiting complete responses were rechallenged with CT-26 parental cells and tumor growth was monitored. Naïve mice challenged with parental CT-26 cells served as controls. G CT-26 parental and Tak1 -deficient tumor growth was monitored in Balb/c mice administered an α-CD8 depleting antibody for two sequential days prior to tumor engraftment and during tumor progression at the indicated days ( n = 10/group). H CT-26 parental and Tak1 -deficient tumor growth was assessed in Balb/c mice administered an α-TNF-α neutralizing antibody (15 mg/kg) at the indicated days ( n = 10/group). Tumor growth curves represent the mean tumor volume +/− SEM. Statistical differences between tumor volumes at the final measurement was determined using a two-way ANOVA with Tukey’s multiple correction where * p < 0.05; ** p < 0.01; and **** p < 0.0001 was considered significant.

Article Snippet: For the CD8 depletion protocol, mice were injected with anti-mouse CD8 (BioXCell; BP0061, New Haven, CT) or IgG2a Isotype control (BioXCell; BP 00085) at 10 mg/kg i.p. at the dose schedule of Day -2, Day -1 prior to flank inoculation of cells, then again on Days 4, 8, and 11 post inoculation.

Techniques: Clone Assay

Journal: Experimental & molecular medicine

Article Title: Neutrophil extracellular traps mediate the crosstalk between plaque microenvironment and unstable carotid plaque formation.

doi: 10.1038/s12276-024-01281-4

Figure Lengend Snippet: Fig. 4 MD-1 induces the formation of NETs. a Isolation and validation of neutrophils (Giemsa staining, Trypan blue staining, and cell counter, scale bar = 20 μm). b Hochest staining and ELISA were used to quantify the concentration and time course of MD-1-induced NETs. c NETs were quantified in the negative control group, the MD-1 (75 ng/ml) group, and the two positive control groups. d Co-IP revealed that MD-1 and TLR4 coexisted on neutrophils. e The interaction between MD-1 and TLR4 in peripheral blood neutrophils from carotid plaque patients and peripheral blood neutrophils from healthy volunteers induced under different conditions was analyzed by Duolink PLA (scale bar = 100 μm). f Western blot analysis showed that neutrophils induced with MD-1 upregulated TLR4 expression and increased the phosphorylation of IRAK1/IRAK4, p38 MAPK, and ERK1/2 in a time-dependent manner (from 0 to 60 min). t total value; p phosphorylation value. β-Actin was used as an internal control.

Article Snippet: We constructed a neutrophil depletion model based on the in vivo injection of InVivoPlus anti-mouse Ly6G (BP0075-1-5MG, BioXCell, New Hampshire, USA).

Techniques: Isolation, Biomarker Discovery, Staining, Enzyme-linked Immunosorbent Assay, Concentration Assay, Negative Control, Positive Control, Co-Immunoprecipitation Assay, Western Blot, Expressing, Phospho-proteomics, Control