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Blood, Vol. 108, Issue 9, 3135-3142, November 1, 2006
Altered B-cell receptor signaling kinetics distinguish human follicular lymphoma B cells from tumor-infiltrating nonmalignant B cells
Blood Irish et al.
108: 3135
Supplemental materials for: Irish et al
Files in this Data Supplement:
- Figure S1. Basal phosphorylation in lymphoma and normal B cells (PDF, 586 KB) -
Flow cytometry analysis of resting phospho-protein levels in triplicate tests of normal PBMC B cells from three donors (PBMC), a lymphoma cell line (Ramos), and FL tumor B cells from three different patients (FL). Differences in median fluorescence intensity (MFI) relative to a control PBMC sample (blue line) have been indicated by coloring the peaks. Black indicates no difference, lighter yellow indicates increased MFI relative to the control, and blue indicates decreased MFI.
- Figure S2. Multidimensional analysis indicates significant BCR mediated Erk1/2 phosphorylation can occur independent of Syk phosphorylation in FL B cells (PDF, 435 KB) -
(A) Flow cytometry contour plots of PBMC and two different FL patient biopsies (FL-P09, FL-P11) compare signaling in two dimensions (p-Erk1/2 vs. p-Syk) in CD20+ B cells stimulated by BCR crosslinking ( -µ-µ/ ) for various times in a short time course (1, 2, 4, 8, or 16 min) or left unstimulated (0 min). Arrows highlight cells where phosphorylation of Syk and Erk1/2 that occurred simultaneously (e.g. PBMC, 2 min) or cells where of Erk1/2 phosphorylation occurred without Syk phosphorylation (e.g. FL-P09, 2 min).
(B) Flow cytometry contour plots of PBMC and two different FL patient biopsies (FL-P09, FL-P11) compare signaling in two dimensions (p-Erk1/2 vs. p-Syk) in CD20+ B cells stimulated by BCR crosslinking plus H2O2 (-µ/ + H2O2) for various times in a short time course (1, 2, 4, 8, or 16 min) or left unstimulated (0 min). Arrows highlight cells the simultaneous per-cell phosphorylation of Syk and Erk1/2 that occurs in all three samples.
- Figure S3. H2O2 stimulates ligand independent signaling in Ramos but not in primary B cells (PDF, 245 KB) -
Flow cytometry analysis of phospho-protein levels in PBMC, FL, and Ramos cells exposed to 3.3 mM H2O2 without BCR crosslinking for various times in a short time course (1, 2, 4, 8, or 16 min) or left unstimulated (0 min).
- Figure S4. Comparison of tumor and non-tumor B cells in FL patient samples stimulated by BCR crosslinking alone (PDF, 213 KB) -
BCR-mediated signaling at 4 and 30 minutes following BCR crosslinking alone was measured in subsets of the CD20+ B cells from four FL patient samples (FL-P07, FL-P 08, FL-P 09, FL-P 11). FL B cells (CD20+ Bcl-2hi non-tumor light chain-, dark arrow) and tumor infiltrating non-malignant B cells (CD20+ Bcl-2lo non-tumor light chain+, light arrow) were distinguished and their signaling compared by coloring heat map squares relative to the fold induction of phosphorylation relative to the unstimulated sample (0 min).
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