Journal: Journal of experimental & clinical cancer research : CR

Article Title: Regnase-1 downregulation promotes pancreatic cancer through myeloid-derived suppressor cell-mediated evasion of anticancer immunity.

doi: 10.1186/s13046-023-02831-w

Figure Lengend Snippet: Fig. 6 Suppression of CTLs by CD11b+ MDSCs is responsible for the acceleration of tumor progression by Regnase-1 downregulation. A-D Evaluation of phenotypes of orthotopic syngeneic tumors of WT or Regnase-1 KO murine pancreatic cancer cells. Representative macro images of pancreatic tumors (A). Relative mRNA levels of Cd8a, Ifng, Fasl, and Gzmb (B) (N = 6 per group). Representative images of HE (C, left panel) and CD8a immunostaining (C, right panel) and the number of CD8-positive cells (C, right) (N = 6 per group). Dot plots of CD3+CD8+ cells evaluated by flow cytometry (D, left) and the proportion of CD8 + cells among CD45+ cells (D, right) (N = 3 per group). E–H Evaluation of phenotypes of orthotopic syngeneic tumors of WT or Regnase-1-KO murine pancreatic cancer cells with or without depletion of CD8+ cells upon anti-CD8a antibody or IgG treatment. Experimental schematic (E). Dot plots of CD3+ and CD8.+ cells in WT or Regnase-1-KO syngeneic tumors upon anti-CD8a antibody or IgG treatment evaluated by flow cytometry (F). Tumor weights (G) (N = 6 per group). The relative mRNA levels of Cd8a, Ifng, Fasl, and Gzmb (H) (N = 6 per group). Student’s t test was used to evaluate differences between 2 groups. One-way ANOVA with Tukey’s post hoc test was used to compare differences among 4 groups. *P < 0.05, scale bars: 100 μm (insets)

Article Snippet: BE0061, a fully neutralizing mAb recognizing CD8a, and control IgG were obtained from Bioxcell.

Techniques: Immunostaining, Flow Cytometry

Journal: Kidney international

Article Title: Leptin induces TGF-beta synthesis through functional leptin receptor expressed by human peritoneal mesothelial cell.

doi: 10.1038/sj.ki.5000409

Figure Lengend Snippet: Figure 10 | Blocking study of TGF-b release by HPMC. Concentration of TGF-b in culture supernatant from HPMC incubated with medium containing 5.6 mM glucose, 40 mM glucose, 20 ng/ml leptin, 40 mM glucose plus 20 ng/ml leptin, or glucose-HPAC conditioned medium in the absence (white bar) presence of anti-leptin receptor antibody (crossed bar; last bar in each group; 50mg/ml), inhibitor to JAK2 (AG490; 10 mM; dotted bar; 2nd bar in each group), PKC (calphostin; 100 nM; gray bar) or both (10 mM AG490 plus 100 nM calphostin; dark bar). Data are mean7s.d. of four individual experiments.

Article Snippet: Monoclonal anti-Ob-Rb, neutralizing anti-leptin receptor antibody and leptin protein were obtained from R&D System (Minneapolis, MN, USA).

Techniques: Blocking Assay, Concentration Assay, Incubation

Journal:

Article Title: Differential regulation of metabolic, neuroendocrine, and immune function by leptin in humans

doi: 10.1073/pnas.0505429103

Figure Lengend Snippet: Leptin depletion or neutralization inhibits polyclonal T cell proliferation and mixed lymphocyte reactions (MLR), respectively. (a–c)The proliferative response of T lymphocytes to polyclonal stimuli (OKT3, PHA, and PMA/Iono) from controls is completely inhibited in medium with human serum depleted of leptin. Addition of recombinant human leptin (at 100 ng/ml final concentration) completely reverses this phenomenon. (d) Anti-leptin blocking Abs partially inhibit the antigen-specific proliferative response of T cells during MLR. HS, human serum.

Article Snippet: For leptin depletion from serum, a protein G-Sepharose affinity column (Amersham Pharmacia) was used after adhesion on G protein of a polyclonal rabbit anti-human leptin Ab (BioVendor).

Techniques: Neutralization, Recombinant, Concentration Assay, Blocking Assay

Journal: eClinicalMedicine

Article Title: Efficacy of a monovalent (D614) SARS-CoV-2 recombinant protein vaccine with AS03 adjuvant in adults: a phase 3, multi-country study

doi: 10.1016/j.eclinm.2023.102168

Figure Lengend Snippet: Variant distribution by country and calendar time in all participants regardless of prior SARS-CoV-2 infection.

Article Snippet: The levels of neutralizing antibodies in serum samples were assessed using a validated SARS-CoV-2 pseudovirus neutralization assay (Monogram Biosciences, South San Francisco, CA, USA), as previously described.

Techniques: Variant Assay, Infection

Journal: European journal of immunology

Article Title: Leptin-mediated cytokine release and migration of eosinophils: implications for immunopathophysiology of allergic inflammation.

doi: 10.1002/eji.200636866

Figure Lengend Snippet: Figure 1. Effects of leptin on viability on eosinophils. Eosinophils (5 105/well) were cultured with or without leptin (50 ng/mL) for 24 h in a 24-well plate. Apoptosis of eosinophils was assessed by the TACSTM Annexin V-FITC assay using flow cytometry. Representative dot plots of (A) medium control (CTL), and (B) leptin-treated eosinophils were shown from triplicate experiments.

Article Snippet: Anti-human leptin neutralization mAb was from R&D Systems (MN, USA) and mouse IgG1 antibody was from BD PharMingen (CA, USA).

Techniques: Cell Culture, Flow Cytometry, Control

Journal: European journal of immunology

Article Title: Leptin-mediated cytokine release and migration of eosinophils: implications for immunopathophysiology of allergic inflammation.

doi: 10.1002/eji.200636866

Figure Lengend Snippet: Figure 2. Effects of leptin on cell surface expression of ICAM-1, CD18, ICAM-3 and L-selectin on eosinophils. Eosinophils (5 105/well) were cultured with leptin (0–100 ng/mL) for 16 h in a 24-well plate. Surface expression of adhesion molecules per 10 000 viable cells was analyzed by flow cytometry as MFI; (A) representative histograms of the changes in the expression of adhesion molecules. Results of (B) ICAM-1, (C) CD18, (D) ICAM-3 and (E) L-selectin have been normalized by subtracting appropriate isotypic control and are expressed as the arithmetic mean SD of five independent experiments. * p<0.05, ** p<0.01 and *** p<0.001 when compared with medium control.

Article Snippet: Anti-human leptin neutralization mAb was from R&D Systems (MN, USA) and mouse IgG1 antibody was from BD PharMingen (CA, USA).

Techniques: Expressing, Cell Culture, Flow Cytometry, Control

Journal: European journal of immunology

Article Title: Leptin-mediated cytokine release and migration of eosinophils: implications for immunopathophysiology of allergic inflammation.

doi: 10.1002/eji.200636866

Figure Lengend Snippet: Fig. 10 shows that leptin-induced release of IL-1b and IL-6, and IL-8, GRO-a and MCP-1 was suppressed by BAY11–7082 and SB203580, and BAY11–7082, PD98059 and SB203580, respectively (all p<0.05).

Article Snippet: Anti-human leptin neutralization mAb was from R&D Systems (MN, USA) and mouse IgG1 antibody was from BD PharMingen (CA, USA).

Techniques:

Journal: European journal of immunology

Article Title: Leptin-mediated cytokine release and migration of eosinophils: implications for immunopathophysiology of allergic inflammation.

doi: 10.1002/eji.200636866

Figure Lengend Snippet: Figure 4. Effects of neutralization antibody against leptin on the biological activities of leptin on eosinophils. Leptin was incubated with or without neutralization antibody (0.1–10 lg/mL) for 15 min and then incubated with eosinophils (5 105/well) for the assay of (A) apoptosis, (B) cell surface of adhesion molecules, (C) chemokinesis and (D) induction of cytokines. Results were expressed as arithmetic mean plus SD from triplicate experiments. Mouse IgG1 was used as a nonspecific antibody for isotypic antibody control. # p<0.05; ## p<0.01 when compared with medium control (CTL); * p<0.05, ** p<0.01, *** p<0.001 when compared with leptin control (lep). Ab0.1: neutralization antibody (0.1 lg/mL); Ab1: neutralization antibody (1 lg/mL); Ab10: neutralization antibody (10 lg/mL); mIgG: mouse IgG1 (10 lg/mL).

Article Snippet: Anti-human leptin neutralization mAb was from R&D Systems (MN, USA) and mouse IgG1 antibody was from BD PharMingen (CA, USA).

Techniques: Neutralization, Incubation, Control

Journal: European journal of immunology

Article Title: Leptin-mediated cytokine release and migration of eosinophils: implications for immunopathophysiology of allergic inflammation.

doi: 10.1002/eji.200636866

Figure Lengend Snippet: Figure 5. Representative expression profiles of intracellular phosphorylation of MAPK and other serine/threonine kinases of eosinophils upon activation by leptin. Eosinophils (1 107/ well) were treated with or without leptin (50 ng/mL) for 15 min. Eosinophils were then washed and lysed. Twenty-one different intracellular phosphorylated MAPK and other serine/threonine kinases in total cellular lysate were assessed semi-quantita- tively using an antibody based Human Phospho-MAPK Array Kit. Positive and negative controls were designated at (A1, A2, A21, A22, F1, F2) and (E3–E14), respectively. Triplicate experi- ments were performed with essentially identical results. The format of the antibodies on the array are detailed in Table 2.

Article Snippet: Anti-human leptin neutralization mAb was from R&D Systems (MN, USA) and mouse IgG1 antibody was from BD PharMingen (CA, USA).

Techniques: Expressing, Phospho-proteomics, Activation Assay

Journal: European journal of immunology

Article Title: Leptin-mediated cytokine release and migration of eosinophils: implications for immunopathophysiology of allergic inflammation.

doi: 10.1002/eji.200636866

Figure Lengend Snippet: Figure 6. Activation of ERK, p38 MAPK, JAK2 and NF-jB activities in eosinophils by leptin. Eosinophils (1 107/well) were cultured with or without leptin (50 ng/mL) for different indicated incubation times. Total cellular proteins were extracted from eosinophils for the measurement of (A) phosphorylated and total p38 MAPK, (B) phosphorylated and total ERK, (C) phosphorylated and total IjBa, and (D) phosphorylated and total JAK2 by Western blot analysis. Experiments were performed in three independent replicates with essentially identical results, and representative results are shown.

Article Snippet: Anti-human leptin neutralization mAb was from R&D Systems (MN, USA) and mouse IgG1 antibody was from BD PharMingen (CA, USA).

Techniques: Activation Assay, Cell Culture, Incubation, Western Blot

Journal: European journal of immunology

Article Title: Leptin-mediated cytokine release and migration of eosinophils: implications for immunopathophysiology of allergic inflammation.

doi: 10.1002/eji.200636866

Figure Lengend Snippet: Figure 7. Effects of AG490, BAY11–7082, LY294002, PD98059, SB203580 and SP600125 on viability of leptin-treated eosino- phils. Eosinophils (5 105/well) were pretreated with AG490 (10 lM), BAY11–7082 (1 lM), PD98059 (10 lM), LY294002 (5 lM), SB203580 (7.5 lM) or SP600125 (3 lM) for 1 h followed by incubation with or without leptin (50 ng/mL) for 24 h in a 24- well plate. Apoptosis of eosinophils was assessed by the TACSTM Annexin V-FITC assay using flow cytometry. Results were expressed as arithmetic mean plus SD from triplicate experiments. ## p<0.01 when compared with medium control; * p<0.05 when compared with the leptin control. AG: AG490; BAY: BAY11–7082; PD: PD98059; LY: LY294002; SB: SB203580; SP: SP600125; DM: DMSO-

Article Snippet: Anti-human leptin neutralization mAb was from R&D Systems (MN, USA) and mouse IgG1 antibody was from BD PharMingen (CA, USA).

Techniques: Incubation, Flow Cytometry, Control

Journal: European journal of immunology

Article Title: Leptin-mediated cytokine release and migration of eosinophils: implications for immunopathophysiology of allergic inflammation.

doi: 10.1002/eji.200636866

Figure Lengend Snippet: Figure 8. Effects of AG490, BAY11–7082, LY294002, PD98059, SB203580 and SP600125 on leptin-induced cell surface expression of (A) ICAM-1, (B) ICAM-3, (C) CD18, and (D) L-selectin on eosinophils. Eosinophils (5 105/well) were pretreated with AG490 (10 lM), BAY11–7082 (1 lM), PD98059 (10 lM), LY294002 (5 lM), SB203580 (7.5 lM) or SP600125 (3 lM) for 1 h followed by incubation with or without leptin (50 ng/mL) for further 16 h. Surface expression of the adhesion molecules of 10 000 cells was assessed by flow cytometry as MFI. Results are expressed as the arithmetic mean plus SD from five independent experiments. DMSO (0.1%) was used as the DMSO control. ### p<0.001 when compared with medium control; * p<0.05, ** p<0.01 when compared with the leptin control.

Article Snippet: Anti-human leptin neutralization mAb was from R&D Systems (MN, USA) and mouse IgG1 antibody was from BD PharMingen (CA, USA).

Techniques: Expressing, Incubation, Flow Cytometry, Control

Journal: European journal of immunology

Article Title: Leptin-mediated cytokine release and migration of eosinophils: implications for immunopathophysiology of allergic inflammation.

doi: 10.1002/eji.200636866

Figure Lengend Snippet: Figure 9. Effects of AG490, BAY11–7082, LY294002, PD98059, SB203580 and SP600125 on the migration of eosinophils induced by leptin. Eosinophils (5 104 cells) were pretreated with AG490 (10 lM), BAY11–7082 (1 lM), PD98059 (10 lM), LY294002 (5 lM), SB203580 (7.5 lM) or SP600125 (3 lM) for 1 h followed by incubation with or without leptin (50 ng/mL) for further 16 h. Eosinophils were then incubated in a 48-well microchamber in the upper chambers, while the RPMI medium was placed in the lower chambers. Migration was carried out for 6 h. Results are expressed as cell number/high power field with arithmetic mean plus SD of three independent experi- ments. # p<0.05 when compared with medium control; * p<0.05 when compared with the leptin control.

Article Snippet: Anti-human leptin neutralization mAb was from R&D Systems (MN, USA) and mouse IgG1 antibody was from BD PharMingen (CA, USA).

Techniques: Migration, Incubation, Control