Journal: Cancer Immunology, Immunotherapy : CII

Article Title: Distinct immune signatures for predicting the immunotherapy efficacy of esophageal squamous cell carcinoma or adenocarcinoma

doi: 10.1007/s00262-024-03904-1

Figure Lengend Snippet: Detecting unique cytokines linked to clinical outcomes in esophageal squamous cell carcinoma and adenocarcinoma separately. a Venn diagram shows genes in the cytokine-cytokine receptor pathway for ESCC and EAC from TCGA, annotated with gene counts in distinct and overlapping categories. b Overall survival of patients with TCGA-ESCC and different expression levels of TNFSF10 and CXCL14. c TNFSF10 and CXCL14 expression comparison between early and advanced ESCC tissues (TCGA). d Survival probabilities of patients with EAC classified by CXCL8 and IL17RB expression levels (TCGA). e CXCL8 and IL17RB expression during EAC progression from early to advanced stages. f TNFSF10 and CXCL14 expression in early and advanced ESCC tissues of Chinese patients. g, h The left panel presents immunohistochemical images illustrating TNFSF10 or CXCL14 staining in tissue microarrays obtained from patients diagnosed with ESCC, arranged sequentially from left to right based on ascending expression levels quantified by the H-score. On the far left, the lowest expression level is shown, followed by a sample in the lowest 33.33 percentile, then a moderate expression sample in the next 33.33 percentile, and finally, the highest expression level on the far right. The right panel displays survival curves stratified by the expression of TNFSF10 and CXCL14, categorized into low and high expression groups based on their H-scores. i, j The left panel displays immunohistochemical staining for CXCL8 or IL17RB in tissue microarrays from patients with EAC, with samples ordered from left to right by increasing H-scores. The sequence starts with the lowest expression level, followed by a sample in the first 33.33 percentile, then a sample with moderate expression in the next percentile, and ends with the highest expression level on the far right. P values were calculated using the Mann–Whitney (log-rank) test. Early stage: Stage 0-II; Advanced stage: Stage III-IV; ns: no significance; * P < 0.05, Mann–Whitney test

Article Snippet: The tissue sections were incubated with primary antibodies against TNFSF10 (Catalog No. ab2056; Abcam), CXCL14 (catalog no. ab137541; Abcam), CXCL8 (catalog no. ab106350; Abcam), IL17RB (Catalog No. AF1207; R&D Systems), CXCL10 (Catalog No. ab306587; Abcam, Cambridge, UK), HIF1α (catalog no. ab51608; Abcam), CSF2 (catalog no. ab316862; Abcam), and NOS2 (catalog no. ab283655; Abcam) (diluted at 1:500 in phosphate-buffered saline) overnight at 4℃.

Techniques: Expressing, Comparison, Immunohistochemical staining, Staining, Sequencing, MANN-WHITNEY

Journal: The Journal of Experimental Medicine

Article Title: IL-25 augments type 2 immune responses by enhancing the expansion and functions of TSLP-DC–activated Th2 memory cells

doi: 10.1084/jem.20070406

Figure Lengend Snippet: Distinctive expression of IL-25R by human Th2 memory cells. (a) Expression of IL-25R was measured by quantitative PCR on a panel of human cDNA templates made from various cell types, as described in Materials and methods. Different T cell subsets were isolated as previously described (reference ). (b) Expression of IL-25R isoforms by activated Th2 memory cells was demonstrated by RT-PCR using primers, as described in Materials and methods. Sorted resting T cell subsets, or activated Th2 memory cells stimulated by autologous TSLP-DCs, the homeostatic cytokine IL-7/15, and anti-CD3/CD28 for 7 d were isolated for cell lysate preparation (c) or flow cytometry analysis (d). Quantified cell lysates were subjected to Western blot analysis using biotinylated goat anti–human IL-17RB polyclonal antibody (c). Surface expression of IL-25R of activated Th2 memory cells or resting T cell subsets were examined using flow cytometry, as described in Materials and methods (d). Shaded histograms represent IL-25 R staining; open histograms represent the isotype control. Relative fold differences in IL-25R gene expression between samples listed in panel a are indicated on the x axis. Data are from one of three independent experiments.

Article Snippet: In some experiments, sorted resting T cell subsets or activated CRTH2 + CD4 + Th2 memory cells were stained with biotinylated goat anti–human IL-17RB polyclonal antibody (R&D Systems) and revealed with streptavidin-PE.

Techniques: Expressing, Real-time Polymerase Chain Reaction, Isolation, Reverse Transcription Polymerase Chain Reaction, Flow Cytometry, Western Blot, Staining, Control, Gene Expression

Journal: The Journal of Experimental Medicine

Article Title: IL-25 augments type 2 immune responses by enhancing the expansion and functions of TSLP-DC–activated Th2 memory cells

doi: 10.1084/jem.20070406

Figure Lengend Snippet: IL-25 enhances the proliferation and Th2 polarization of stimulated Th2 memory cells. Sorted Th2 memory T cells were cultured with the cytokines IL-7 only, IL-15 plus IL-7, anti-CD3/CD28, or autologous CD11c + DCs activated by various stimuli (x axis) at a DC/T cell ratio of 1:2 for 7 d in the presence or absence of IL-25. The neutralizing recombinant protein IL-17RB-Fc was used in culture. Their proliferative responses were compared, and graphic bars represent the times of expansion by dividing the final cell number by the initial T cell number in six individual experiments (a). To characterize the effect of IL-25 on Th2 memory cells, sorted cells were cultured in medium only or expanded by IL-15 plus IL-7, anti-CD3/CD28, or TSLP-DCs for 7 d in the presence or absence of IL-25. The proliferated cells were collected and restimulated with PMA plus ionomycin for analysis of intracellular cytokine production (b), or Th2 memory cells expanded by TSLP-DCs in the presence or absence of IL-25 for 7 d were collected and restimulated with anti-CD3/CD28 for 24 h before the measurements of cytokines in the culture supernatants by ELISA (c). For phenotypic analyses, sorted Th2 memory cells cultured in IL-25 alone, or expanded by TSLP-DCs, or TSLP-DCs plus IL-25 were examined using flow cytometry (d). Shaded histograms represent staining of Th2 memory T cells with the markers indicated below histograms; open histograms represent the isotype control. Numbers within the quadrants indicate the percentage of expanded cells that stained positive for each respective cytokine. The results in panels a, b, c, and d are for separate experiments. Data represent the mean ± SD of five experiments.

Article Snippet: In some experiments, sorted resting T cell subsets or activated CRTH2 + CD4 + Th2 memory cells were stained with biotinylated goat anti–human IL-17RB polyclonal antibody (R&D Systems) and revealed with streptavidin-PE.

Techniques: Cell Culture, Recombinant, Enzyme-linked Immunosorbent Assay, Flow Cytometry, Staining, Control

Journal: The Journal of Experimental Medicine

Article Title: IL-25 augments type 2 immune responses by enhancing the expansion and functions of TSLP-DC–activated Th2 memory cells

doi: 10.1084/jem.20070406

Figure Lengend Snippet: IL-25 induces elevated cytokine production by TSLP-DC–activated Th2 memory cells. Freshly isolated resting or activated Th2 memory cells expanded by TSLP-DCs for 7 d were collected, washed, and stimulated directly with IL-25 or cultured in medium only for 24 h in the presence or absence of neutralizing soluble receptor IL-17RB-FC or anti-IL-4 mAbs. Cultured supernatants were collected for ELISA analyses. Data represent the mean ± SD of five experiments.

Article Snippet: In some experiments, sorted resting T cell subsets or activated CRTH2 + CD4 + Th2 memory cells were stained with biotinylated goat anti–human IL-17RB polyclonal antibody (R&D Systems) and revealed with streptavidin-PE.

Techniques: Isolation, Cell Culture, Enzyme-linked Immunosorbent Assay

Journal: The Journal of Experimental Medicine

Article Title: IL-25 augments type 2 immune responses by enhancing the expansion and functions of TSLP-DC–activated Th2 memory cells

doi: 10.1084/jem.20070406

Figure Lengend Snippet: Eosinophil-secreted IL-25 promotes cytokine production by TSLP-DC–activated Th2 memory cells. Resting or TSLP-DC–activated Th2 memory cells purified from normal (a and b) or atopic subjects (a) were cultured in medium only or co-cultured with autologous eosinophils for 3 d in the presence or absence of anti–IL-4 Abs or IL-17RB-Fc recombinant proteins (a). Co-cultured supernatants were harvested for the measurement of Th2 cytokine production using ELISA analyses (a). Activated Th2 memory cells cultured in medium alone or co-cultured with eosinophils for 3 d were collected and restimulated with PMA plus ionomycin before being stained with CD4 and the indicated intracellular cytokines. Numbers within the quadrants indicate the percentage of cells that stained positive for each respective cytokine. Data are from five normal or four atopic subjects and represent the mean ± SD of tested samples.

Article Snippet: In some experiments, sorted resting T cell subsets or activated CRTH2 + CD4 + Th2 memory cells were stained with biotinylated goat anti–human IL-17RB polyclonal antibody (R&D Systems) and revealed with streptavidin-PE.

Techniques: Purification, Cell Culture, Recombinant, Enzyme-linked Immunosorbent Assay, Staining