|
|
Blood, Vol. 110, Issue 5, 1603-1606, September 1, 2007
Deficient CD4+ CD25+ FOXP3+ T regulatory cells in acquired aplastic anemia
Blood Solomou et al.
110: 1603
Supplemental materials for Solomou et al, Vol.110, Issue 5, 1603-1606
Files in this Data Supplement:
- Document 1. Supplemental methods (PDF, 19.1 KB)
- Table S1. Patient demographics (PDF, 32.1 KB) -
CBC indicates complete blood count; Tx, treatment; Hgb, hemoglobin; PLT, platelets; F, female; M, male; SAA, severe aplastic anemia; ATG/CsA, antithymocyte globulin and cyclosporine; N/A, data not available.
- Figure S1. Phenotypic analysis of magnetic-assisted cell sorting (MACS)-purified CD4+CD25− and CD4+ CD25+ T cells from a patient (JPG, 85.4 KB)
-
From left to right: Isotype control, CD4 FITC and CD25PE staining of PBMCs (upper row), of purified CD4+ CD25−, and CD4+ CD25+ T cells (second row).
- Figure S2. Cytoplasmic and nuclear extracts from CD4+ CD25+ T cells from patients and healthy donors were analyzed by immunoblot for NFAT1 and FOXP3 expression, respectively (JPG, 49.6 KB)
-
All patients examined showed decreased or undetectable NFAT1 and FOXP3 protein levels compared to healthy controls.
- Figure S3. Purified CD4+ CD25− and CD4+ CD25− T cells from patients and healthy donors were analyzed by confocal microscopy for FOXP3 expression after transient transfection with a wild-type NFAT1 construct (JPG, 96 KB)
-
CD4+ CD25+ T cells from healthy donors did not show any difference in FOXP3 expression after transfection with the NFAT1 construct; CD4+ CD25− T cells from healthy donors and from patients with aplastic anemia showed aberrant FOXP3 expression after the transfection. CD4+ CD25+ T cells from patients also showed significantly increased FOXP3 expression after the transfection. Patients’ and controls’ cells were transfected and stained in parallel. The 2-D projections (using maximum-pixel-intensity algorithm) of the 3-D fluorescence were generated and the FOXP3 fluorescence intensities are color-coded using a look-up-table with a 0 to 4095 range to create pseudocolor-mapped images. The graphs show the integrated sum of total FOXP3 fluorescence through all optical sections as calculated in the 3-D series. Representative results are shown from at least 4 different experiments.
|
|
