Journal: Scientific Reports

Article Title: Soluble interleukin-27 receptor alpha is a valuable prognostic biomarker for acute graft-versus-host disease after allogeneic haematopoietic stem cell transplantation

doi: 10.1038/s41598-018-28614-4

Figure Lengend Snippet: The expression of sIL-27Rα level in GVHD patients. ( A ) Serum sIL-27Rα levels were significantly increased on the day of neutrophil engraftment compared with pre-conditioning ( P < 0.0001). ( B ) The serum sIL-27Rα levels in patients with grade II–IV aGVHD were significantly lower than those of 0-I aGVHD patients on the day of neutrophil engraftment ( P < 0.001). ( C ) On the day of neutrophil engraftment, serum sIL-27Rα levels in patients with skin aGVHD ( P < 0.01) and liver aGVHD ( P < 0.05) were significantly lower than those of 0-I aGVHD patients. ( D ) There were no similar significant difference regarding cGVHD. Data shown are mean ± SD.

Article Snippet: After washing, biotinylated goat anti-human IL-27Rα polyclonal antibody (100 μL/well, Catalogue #BAF1479, R&D Systems) was added as detection antibody for 2 h at room temperature.

Techniques: Expressing

Journal: Scientific Reports

Article Title: Soluble interleukin-27 receptor alpha is a valuable prognostic biomarker for acute graft-versus-host disease after allogeneic haematopoietic stem cell transplantation

doi: 10.1038/s41598-018-28614-4

Figure Lengend Snippet: The diagnostic value of sIL-27Rα in aGVHD and the association of sIL-27Rα with aGVHD severity, relapse and survival. ( A ) The area under the ROC curve (AUC) was 0.735 (95% CI 0.618–0.853, P = 0.001) on the day of neutrophil engraftment; when using 59.4 ng/ml as cut-off value from the ROC curve, the sensitivity and specificity were 56% and 81%, respectively. ( B ) The cumulative incidence of grade II–IV aGVHD was significantly lower in patients with high sIL-27Rα levels by Gray’s test ( P = 0.004). ( C ) Patients with high sIL-27Rα levels showed favourable overall survival compared with patients with low sIL-27Rα levels with Kaplan-Meier survival analysis by log rank test ( P < 0.001). ( D , E ) Patients with high sIL-27Rα levels had lower relapse rate (CIR) and non-relapse mortality (NRM) than did patients with low sIL-27Rα levels by Gray’s test (P = 0.008, respectively).

Article Snippet: After washing, biotinylated goat anti-human IL-27Rα polyclonal antibody (100 μL/well, Catalogue #BAF1479, R&D Systems) was added as detection antibody for 2 h at room temperature.

Techniques: Diagnostic Assay

Journal: Scientific Reports

Article Title: Soluble interleukin-27 receptor alpha is a valuable prognostic biomarker for acute graft-versus-host disease after allogeneic haematopoietic stem cell transplantation

doi: 10.1038/s41598-018-28614-4

Figure Lengend Snippet: The association of sIL-27Rα levels at neutrophil engraftment with clinical factors.

Article Snippet: After washing, biotinylated goat anti-human IL-27Rα polyclonal antibody (100 μL/well, Catalogue #BAF1479, R&D Systems) was added as detection antibody for 2 h at room temperature.

Techniques: Biomarker Discovery

Journal: Scientific Reports

Article Title: Soluble interleukin-27 receptor alpha is a valuable prognostic biomarker for acute graft-versus-host disease after allogeneic haematopoietic stem cell transplantation

doi: 10.1038/s41598-018-28614-4

Figure Lengend Snippet: Univariate and multivariate analyses of factors affecting the incidence of grade II–IV acute graft- versus-host disease after allogeneic hematopoietic stem cell transplantation at neutrophil engraftment.

Article Snippet: After washing, biotinylated goat anti-human IL-27Rα polyclonal antibody (100 μL/well, Catalogue #BAF1479, R&D Systems) was added as detection antibody for 2 h at room temperature.

Techniques: Transplantation Assay

Journal: Scientific Reports

Article Title: Soluble interleukin-27 receptor alpha is a valuable prognostic biomarker for acute graft-versus-host disease after allogeneic haematopoietic stem cell transplantation

doi: 10.1038/s41598-018-28614-4

Figure Lengend Snippet: sIL-27Rα as an independent prognostic aGVHD biomarker validated in an independent cohort of 80 patients. ( A ) The area under the ROC curve (AUC) was 0.790 (95% CI 0.688–0.892, P < 0.001) on the day of neutrophil engraftment, using 59.4 ng/ml as the cut-off value. ( B ) The cumulative incidence of grade II–IV aGVHD was significantly lower in patients with high sIL-27Rα levels by Gray’s test ( P < 0.001). ( C ) Patients with high sIL-27Rα levels showed favourable overall survival compared with patients with low sIL-27Rα levels on Kaplan-Meier survival analysis by log rank test ( P = 0.012). ( D , E ) Patients with high sIL-27Rα levels had similar relapse rates (CIR) and lower non-relapse mortality (NRM) than did patients with low sIL-27Rα levels by Gray’s test ( P = 0.0931, P = 0.005, respectively).

Article Snippet: After washing, biotinylated goat anti-human IL-27Rα polyclonal antibody (100 μL/well, Catalogue #BAF1479, R&D Systems) was added as detection antibody for 2 h at room temperature.

Techniques: Biomarker Discovery

Journal: Scientific Reports

Article Title: Soluble interleukin-27 receptor alpha is a valuable prognostic biomarker for acute graft-versus-host disease after allogeneic haematopoietic stem cell transplantation

doi: 10.1038/s41598-018-28614-4

Figure Lengend Snippet: Association of sIL-27Rα with serum cytokine levels in aGVHD. The associations between sIL-27Rα levels with serum IL-27, IL-10, HGF, TNFR1, elafin, REG3α and ST2 levels were analysed by using Spearman’s rank correlation coefficient.

Article Snippet: After washing, biotinylated goat anti-human IL-27Rα polyclonal antibody (100 μL/well, Catalogue #BAF1479, R&D Systems) was added as detection antibody for 2 h at room temperature.

Techniques:

Journal: EMBO Molecular Medicine

Article Title: Activation of IL-27 signalling promotes development of postinfluenza pneumococcal pneumonia

doi: 10.1002/emmm.201302890

Figure Lengend Snippet: Influenza-infected IL-27R-deficient mice were resistant to secondary pneumococcal pneumonia. A Lungs from indicated groups of mice were harvested at the designated time points for assessment of IL-27 by ELISA ( n = 5). B Lung IL-27 production in the groups of mice at 24, 48, 72 h after influenza infection, S. pneumoniae infection or secondary pneumococcal infection following influenza infection ( n = 5). C WT splenocytes or LEC were stimulated with influenza virus (MOI = 1) for the indicated time points. IFN-β, EBI3, and IL-27p28 gene transcript level was detected by quantitative PCR ( n = 3). D IL-27 production in the lungs of IFNAR-deficient or WT mice after primary influenza infection or secondary pneumococcal infection ( n = 5). E Pulmonary pneumococcal burdens in IL-27R-deficient and WT mice at 24 or 48 h following primary pneumococcal infection or secondary pneumococcal infection ( n = 12). F Incidence of bacteraemia was measured in IL-27R-deficient and WT mice at 24 or 48 h following primary pneumococcal infection or secondary pneumococcal infection ( n = 12). G Survival for indicated groups of mice following secondary pneumococcal infection ( n = 12). H Pulmonary pneumococcal burdens in WT mice treated with anti-IL-27 blocking antibodies at 48 h after secondary pneumococcal infection ( n = 12). I Survival for indicated groups of mice treated with anti-IL-27 neutralizing antibodies or isotypical IgG after secondary pneumococcal infection ( n = 12). * p < 0.05, ** p < 0.01, *** p < 0.001 when compared between groups denoted by horizontal lines; # p < 0.05 when compared between indicated groups of mice.

Article Snippet: Recombinant murine or human IL-4, IL-17A, IFN-β, GM-CSF, IL-23, IL-27, anti-mouse IL-17A and anti-mouse IL-27p28 antibodies were purchased from R&D Systems (Minneapolis, MN).

Techniques: Infection, Enzyme-linked Immunosorbent Assay, Virus, Real-time Polymerase Chain Reaction, Blocking Assay

Journal: EMBO Molecular Medicine

Article Title: Activation of IL-27 signalling promotes development of postinfluenza pneumococcal pneumonia

doi: 10.1002/emmm.201302890

Figure Lengend Snippet: IL-27 negatively regulated IL-17A production by γ´ T cell upon pneumococcal infection in mice. A IL-17A production in the lungs from IL-27R-deficient and WT mice at 24 h after secondary pneumococcal challenge following influenza infection ( n = 5). B γδ T cells, CD4+ T cells, CD8+ T cells and NKT cells from IL-27R-deficient and WT mice were purified by cell sorting at 24 h after secondary pneumococcal infection. IL-17A gene expression was measured from different cell types ( n = 5). C Percentages of IL-17A producers among γδ T cells in the lungs from IL-27R-deficient and WT mice at indicated times following secondary pneumococcal challenge ( n = 5). D Lung neutrophil numbers in IL-27R-deficient and WT mice at 24 h following secondary pneumococcal challenge ( n = 5). E Lung MPO activity in IL-27R-deficient and WT mice at 24 h following secondary pneumococcal challenge ( n = 5). F Schematic representation model of γδ T cells adoptive transfer experiment. G Lung IL-17A levels were determined at 24 h after secondary pneumococcal infection ( n = 5). H Lung neutrophil numbers in WT mice transferred with IL-27R-deficient or WT γδ T cells at 24 h after secondary pneumococcal infection ( n = 5). I Lung MPO activity at 24 h after secondary pneumococcal infection ( n = 5). J Pulmonary pneumococcal burdens at 48 h after secondary pneumococcal infection ( n = 12). K Survival for WT mice transferred with IL-27R-deficient or WT γδ T cells after secondary pneumococcal infection ( n = 12). * p < 0.05, ** p < 0.01, *** p < 0.001 when compared between groups denoted by horizontal lines; # p < 0.05 when compared with mice transferred with WT γδ T cells.

Article Snippet: Recombinant murine or human IL-4, IL-17A, IFN-β, GM-CSF, IL-23, IL-27, anti-mouse IL-17A and anti-mouse IL-27p28 antibodies were purchased from R&D Systems (Minneapolis, MN).

Techniques: Infection, Purification, FACS, Expressing, Activity Assay, Adoptive Transfer Assay

Journal: EMBO Molecular Medicine

Article Title: Activation of IL-27 signalling promotes development of postinfluenza pneumococcal pneumonia

doi: 10.1002/emmm.201302890

Figure Lengend Snippet: IL-27 mediated the inhibitory effects of IFNAR signalling on IL-17A production by γ´ T cells during secondary pneumococcal infection. Recombinant murine IL-27 was given i.t. into IFNAR-deficient or WT mice 24 h before intranasal pneumococcal administration, lungs were then collected for analysis at different time points. A Lung IL-17A levels were determined at 24 h following secondary pneumococcal infection ( n = 4). B Percentages of IL-17A producers among γδ T cells in the lungs from groups of mice at indicated times following secondary pneumococcal challenge ( n = 4). C Lung neutrophil numbers at 24 h after secondary pneumococcal infection ( n = 4). D Lung MPO activity at 24 h after secondary pneumococcal infection ( n = 4). E Pulmonary pneumococcal burdens at 48 h after secondary pneumococcal infection ( n = 8). F Survival for IFNAR-deficient or WT mice treated with or without exogenous IL-27 after secondary pneumococcal infection ( n = 8). * p < 0.05, ** p < 0.01, *** p < 0.001 when compared between IFNAR-deficient mice treated with and without IL-27; # p < 0.05 when compared with mice treated with IL-27.

Article Snippet: Recombinant murine or human IL-4, IL-17A, IFN-β, GM-CSF, IL-23, IL-27, anti-mouse IL-17A and anti-mouse IL-27p28 antibodies were purchased from R&D Systems (Minneapolis, MN).

Techniques: Infection, Recombinant, Activity Assay

Journal: EMBO Molecular Medicine

Article Title: Activation of IL-27 signalling promotes development of postinfluenza pneumococcal pneumonia

doi: 10.1002/emmm.201302890

Figure Lengend Snippet: IL-27 inhibited IL-17A production by γ´ T cells in vitro . A FACS analysis of IL-17A and IFN-γ expression in FACS-sorted lung or spleen γδ T cells cultured for 72 h under the stimulation of HkSp (1 × 10 8 CFU/ml) and IL-23 (50 ng/ml) in the presence or absence of IL-27 (100 ng/ml). B IL-17A concentrations in the supernatants of lung or spleen γδ T cells activated by HkSp and IL-23 in the presence or absence of IL-27 as determined by ELISA. C FACS analysis of IL-17A and IFN-γ expression in FACS-sorted lung γδ T cells from IL-27R-deficient or WT mice, which were co-cultured with BMDC from IL-27R-deficient or WT mice activated by HkSp (1 × 10 8 CFU/ml) in the presence or absence of IL-27 (100 ng/ml) for 72 h. Lung γδ T cells were gated for FACS analysis. D IL-17A concentrations in the supernatants of lung γδ T cells from IL-27R-deficient or WT mice, which were co-cultured with BMDC from IL-27R-deficient or WT mice activated by HkSp in the presence or absence of IL-27 for 72 h. Results were from three independent experiments, and each was performed with cells isolated from three mice. * p < 0.05, *** p < 0.001 when compared between groups denoted by horizontal lines.

Article Snippet: Recombinant murine or human IL-4, IL-17A, IFN-β, GM-CSF, IL-23, IL-27, anti-mouse IL-17A and anti-mouse IL-27p28 antibodies were purchased from R&D Systems (Minneapolis, MN).

Techniques: In Vitro, Expressing, Cell Culture, Enzyme-linked Immunosorbent Assay, Isolation

Journal: EMBO Molecular Medicine

Article Title: Activation of IL-27 signalling promotes development of postinfluenza pneumococcal pneumonia

doi: 10.1002/emmm.201302890

Figure Lengend Snippet: IL-27 directly contributed to IFN-b-mediated inhibition of IL-17A production in γ´ T cells. A, B Spleen γδ T cells isolated from WT mice were restimulated with HkSp (1 × 10 8 CFU/ml) and IL-23 (50 ng/ml) for 72 h in the presence or absence of supernatants from IFN-β-treated splenocytes plus anti-IL-27 neutralizing antibodies or control IgG. IL-17 levels were determined by (A) FACS analysis and (B) ELISA, respectively. C, D WT or IFNAR-deficient spleen γδ T cells were co-cultured with WT or IFNAR-deficient BMDC activated by HkSp (1 × 10 8 CFU/ml) in the presence or absence of IL-27 (100 ng/ml) for 72 h. (C) The percentage of IL-17A-producing γδ T cells was measured by FACS analysis, (D) while IL-17A secretion in the culture supernatants was detected by ELISA. Results were from three independent experiments, and each was performed with cells isolated from three mice. *** p < 0.001 when compared between groups denoted by horizontal lines.

Article Snippet: Recombinant murine or human IL-4, IL-17A, IFN-β, GM-CSF, IL-23, IL-27, anti-mouse IL-17A and anti-mouse IL-27p28 antibodies were purchased from R&D Systems (Minneapolis, MN).

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

Journal: EMBO Molecular Medicine

Article Title: Activation of IL-27 signalling promotes development of postinfluenza pneumococcal pneumonia

doi: 10.1002/emmm.201302890

Figure Lengend Snippet: STAT1 was critical for suppression of IL-17A in γ´ T cells by IL-27 in vitro and in vivo . A Splenocytes were pretreated with JI-1 (1 nM) for 1 h, followed by stimulation with IL-27 (100 ng/ml) for a further 15 min. Phospho-Jak2 and phosphor-Tyk2 were detected by Western blot. B FACS analysis of percentages of IL-17A producers among γδ T cells in FACS-sorted spleen γδ T cells co-cultured with BMDC infected with HkSp (1 × 10 8 CFU/ml) in the presence or absence of IL-27 (100 ng/ml) and of JI-1 (1 nM) for 72 h, and IL-17A concentrations in the culture supernatants were also determined by ELISA. C Splenocytes were pretreated with fludarabine (50 μM) or S31-201 (10 μM) for 1 h, followed by stimulation with IL-27 (100 ng/ml) for a further 15 min. Phospho-STAT1 (Tyr 701) and phosphor-STAT3 (Tyr 705) were detected by Western blot. D FACS analysis of percentages of IL-17A producers among γδ T cells in FACS-sorted spleen γδ T cells co-cultured with BMDC infected with HkSp (1 × 10 8 CFU/ml) in the presence or absence of IL-27 (100 ng/ml) and of fludarabine (50 μM) or S31-201 (10 μM) for 72 h, and IL-17A concentrations in the culture supernatants were also determined by ELISA. E FACS analysis of IL-17A expression in FACS-sorted spleen γδ T cells from STAT1-deficient or WT mice, which were co-cultured with BMDC from STAT1-deficient or WT mice infected by HkSp in the presence or absence of IL-27 for 72 h. The percentages of IL-17A-producing γδ T cells were determined by FACS analysis, while IL-17A concentrations were determined by ELISA. F FACS-sorted spleen γδ T cells were stimulated with HkSp (1 × 10 8 CFU/ml) and IL-23 (50 ng/ml) in the presence or absence of IL-27 (100 ng/ml) for 72 h. Cell lysates were immunoprecipitated either with anti-STAT1 or isotype IgG. Bound DNA was analyzed by PCR with IL-17A promoter site-specific primers. G Eluted DNA was quantitated by quantitative PCR with primers specific for the IL-17A promoter. H Lung IL-17A concentrations in WT mice transferred with STAT1-deficient or WT γδ T cells at 24 h after secondary pneumococcal infection ( n = 5). I Lung neutrophil numbers in WT mice transferred with STAT1-deficient or WT γδ T cells at 24 h after secondary pneumococcal infection ( n = 5). J Lung MPO activity at 24 h after secondary pneumococcal infection ( n = 5). K Pulmonary pneumococcal burdens at 48 h after secondary pneumococcal infection ( n = 12). L Survival for WT mice transferred with STAT1-deficient or WT γδ T cells after secondary pneumococcal infection ( n = 12). * p < 0.05, ** p < 0.01, *** p < 0.001 when compared between groups denoted by horizontal lines; # p < 0.05 when compared with mice transferred with WT γδ T cells.

Article Snippet: Recombinant murine or human IL-4, IL-17A, IFN-β, GM-CSF, IL-23, IL-27, anti-mouse IL-17A and anti-mouse IL-27p28 antibodies were purchased from R&D Systems (Minneapolis, MN).

Techniques: In Vitro, In Vivo, Western Blot, Cell Culture, Infection, Enzyme-linked Immunosorbent Assay, Expressing, Immunoprecipitation, Real-time Polymerase Chain Reaction, Activity Assay

Journal: EMBO Molecular Medicine

Article Title: Activation of IL-27 signalling promotes development of postinfluenza pneumococcal pneumonia

doi: 10.1002/emmm.201302890

Figure Lengend Snippet: IL-27 inhibited IL-17A production in human Vγ9V´2 T cells. A ELISA analysis for IL-27 levels in BAL and serum samples from healthy individuals and influenza-infected patients. B ELISA analysis for IL-27 production in human cells. Human monocyte-derived DC, momocytes, pulmonary epithelial cells and lymphocytes were stimulated with influenza virus (MOI = 1) and HkSp (1 × 10 8 CFU/ml). After 24 h, ELISA was performed to measure the IL-27 concentrations in the culture supernatants. C FACS analysis for IL-17A and IFN-γ expression in human Vγ9Vδ2 T cells co-cultured with DC activated by HkSp (1 × 10 8 CFU/ml) in the presence or absence of IL-27 (100 ng/ml) at 5 days. D IL-17A concentrations in the supernatants of human Vγ9Vδ2 T cells co-cultured with DC activated by HkSp in the presence or absence of IL-27 at 5 days. E The expression of RORγt, IL-23R and CCR6 in human Vγ9Vδ2 T cells co-cultured with DC infected with HkSp in the presence or absence of IL-27 F ELISA analysis for cytokine production from human DC activated by HkSp in the presence or absence of IL-27. Results were from three independent experiments, and each was performed with cells isolated from three different donors. *** p < 0.001 when compared between groups denoted by horizontal lines.

Article Snippet: Recombinant murine or human IL-4, IL-17A, IFN-β, GM-CSF, IL-23, IL-27, anti-mouse IL-17A and anti-mouse IL-27p28 antibodies were purchased from R&D Systems (Minneapolis, MN).

Techniques: Enzyme-linked Immunosorbent Assay, Infection, Derivative Assay, Virus, Expressing, Cell Culture, Isolation