Journal: The Journal of Experimental Medicine
Article Title: Coexpression of CD25 and CD27 identifies FoxP3+ regulatory T cells in inflamed synovia
Figure Lengend Snippet: CD4 + CD25 + T cells are enriched in synovial fluid of JIA patients, express high amounts of FoxP3 mRNA, and efficiently suppress proliferation of CD4 + CD25 − autologous T cells. (A) Mononuclear cells from peripheral blood (PBMC) and synovial fluid (SFMC) were obtained from the same JIA patient and stained with antibodies to CD4 and CD25. Shown is the percentage of CD25 + and CD25 − cells within the CD4 + population. (B) Percentages of CD4 + CD25 + in PBMCs and SFMCs in 15 JIA patients. p-value determined by Wilcoxon rank test. (C) PBMCs and SFMCs were sorted based on expression of CD4 and CD25, and analyzed for expression of FoxP3 mRNA relative to 18S rRNA by quantitative real-time PCR. 1 representative experiment out of 10 is shown. (D) Proliferation of 1.5 × 10 4 CFSE-labeled CD4 + CD25 − peripheral blood T cells stimulated by anti-CD3 and DCs in the presence of equal numbers of autologous CD4 + CD25 − T cells (unshaded histograms) or CD4 + CD25 + T cells (shaded histograms) isolated from PBMCs or SFMCs. The total number of responder T cells that had performed one or more cell divisions was determined by CFSE dilution analysis. Responder T cells proliferated to a similar extent in the absence of CD4 + CD25 − control T cells (not depicted). The percentage inhibition was calculated from the number of dividing responder cells in presence of CD4 + CD25 + T cells as compared with their number in presence of CD4 + CD25 − control cells. One representative experiment out of five is shown.
Article Snippet: Paraffin serial sections were stained for 30 min at room temperature with mouse antibodies to CD4 (4B12), CD25 (25C04), CD27 (137B4; obtained from Neomarkers), CD20 (L26), and CD3 (polyclonal antisera; obtained from DakoCytomation) followed by anti–mouse Ig antibody conjugated to peroxidase-labeled dextran polymer (EnVision; DakoCytomation) and chromogenic diaminobenzidine substrate (DakoCytomation).
Techniques: Staining, Expressing, Real-time Polymerase Chain Reaction, Labeling, Isolation, Inhibition