|
|
Blood, Vol. 111, Issue 7, 3644-3652, April 1, 2008
CD38 induces differentiation of immature transitional 2 B lymphocytes in the spleen
Blood Rodríguez-Alba et al.
111: 3644
Supplementary materials for: Rodríguez-Alba et al
Files in this Data Supplement:
- Figure S1. T2 B cells are larger and more granular than T1 or M B lymphocytes (JPG, 173 KB)
-
Total splenocytes were stained with CD21, CD24 and B220 (as described in the Figure 1). The cells were analyzed considering their forward and side scatter values. The expression of CD21 and CD24 was evaluated within B220 positive cells to quantify T1, T2, and M B lymphocytes. The numbers indicate the percentage of each subset and the dot plots are representative from three independent experiments with similar results.
- Figure S2. IL-4 increased the proliferation of T2 and M B cells induced by anti-CD38 stimulation, whereas T1 subset did not respond (JPG, 93 KB)
-
Total splenic cells were gated on B220 positive cells (100%) and the percentage of each subset was calculated according to the expression of CD21 and CD24. (A) More than 90 % pure Transitional 1 (CD24high and CD21low), Transitional 2 (CD24high and CD21high) or mature follicular (CD24low and CD21intermediate) B cells were used in each experiment. (B) T1, T2 and M B cells were stimulated, in the presence of IL-4, with: rat-IgG (open bars), anti-CD38 (black bars) or anti-IgM (gray bars). The plates were incubated 48 h at 37°C and pulsed with 1 µCi/well of 3H thymidine 8 h before being harvested. The results are expressed as mean and standard deviation from three independent experiments where t-Student test was p<0.01 as indicated.
- Figure S3. Anti-CD38 did not induce T1 B cell differentiation (JPG, 169 KB)
-
Purified T1, T2 and M subsets were activated as indicated in the Figure 2. The cells were harvested at 24 or 48 h and re-stained to evaluate differentiation. (A) Differentiation of T2 B cells in the presence of IL-4, comparing cells stimulated with medium alone, anti-CD38 or anti-IgM. (B) T1 and M B cells at 48h after incubation with the indicated stimuli. The numbers in each dot plot indicate the percentage of cells in each region. Dot plots are representative from three independent experiments with similar results.
- Figure S4. CD38 stimulation induced differentiation of highly purified T2 B lymphocytes (JPG, 64.7 KB)
-
Total splenocytes were stained with B220, CD21, CD24 and CD23, to separate T2 B from MZ B lymphocytes. (A) Numbers of 97% purified T2 B lymphocytes were obtained. T2 B cells were activated and cultured 48 h as indicated in the Figure 3, with or without the addition of IL-4. (B) After incubation, they were re-stained to evaluate their differentiation. The numbers indicate the percentage of each subpopulation. Dot plots are representative from three independent experiments.
- Figure S5. Anti-CD38 stimulation promotes apoptosis of T1 B lymphocytes (JPG, 55.6 KB)
-
Purified T1 B cells were stimulated 12 or 24 h with: medium, anti-CD38 or anti-IgM. After incubation, T1 B cells were permeabilized and stained with Propidium Iodide . The histograms show the content of DNA for each condition. (A) T1 B lymphocytes in the SubG0 region are indicated in each histogram. (B) Percentage of T1 B cell in SubG0 region after 24 h of stimulation with: medium (open bars), anti-CD38 (black bars) or anti-IgM (gray bars). The results are expressed as mean and standard deviation from three independent experiments where t-Student test was: p<0.01 as indicated.
|
|
