LEGEND TO FIGURES
Figure 1. Effect of CD8 Treg (CD183+CCR7+CD45RA-CD8+) on B cell proliferation. Purified CFSE labeled B cells were co-cultured in the presence or absence of various concentrations of CD183+CCR7+CD45RA- CD8 Treg and activated with anti-CD40 and CpG for 5 days. Cell division was measured by flow cytometry. (A) shows a representative flow cytometry histogram. (B) shows cumulative data from 6 different normal donors as mean ± sem. Inhibition was observed only at 1:1 B:CD8 Treg ratio.
Figure 2. Effect of CD8 Treg (CD183+CCR7+CD45RA-CD8+) on immunoglobulin production. Purified B cells were co-cultured in the presence or absence of CD183+CCR7+CD45RA- CD8 Treg at 1:1 ratio and activated with anti-CD40 and CpG for 14 days. Supernatants were collected and immunoglobulins were measured by ELISA assay. Data are expressed for 6 subjects as mean ± sem. CD8 Treg inhibited significantly IgM ( P<0.005) and IgG (P< 0.039 ) production. No significant effect was observed on IgA production (P>0.7).
Figure 3. Further characterization of CD8 Treg. Activated CD8+ T cells were stained with various monoclonal antibodies and isotype controls. FoxP3+ cells were gated and examined for the expression of different antigens. (A) expression of molecules on % of CD8+ T cells, and (B) mean flourescence intensity (MFI) of the different molecules. Data are shown for 5 different subjects and expressed as mean ± sem. A significant (*) increased (<P.005) expression of CD25, CD183, CD278, CTLA-4 , and LAP-1was observed on FoxP3+ CD8+ T cells as compared to FoxP3- cells.
Figure 4. Effect of CD8 Treg (CD183+CD25highCD278+CD8+) on B cell proliferation.Purified CFSE labeled B cells were co-cultured in the presence or absence of various concentrations of CD183+CD25highCD278+ CD8 Treg and activated with anti-CD40 and CpG for 5 days. Cell division was measured by flow cytometry. (A) shows a representative flow cytometry histogram.(B) shows cumulative data from 6 different normal donors as mean ± sem. In a concentration-dependent manner CD8 Treg inhibited B cell proliferation.
Figure 5. Effect of CD8 Treg (CD183+CD25highCD278+CD8+) on immunoglobulin production. Purified B cells were co-cultured in the presence or absence of CD183+CD25highCD278+ CD8 Treg at various ratios and activated with anti-CD40 and CpG for 14 days. Supernatants were collected and immunoglobulins were measured by ELISA assay. Data are shown from 5 separate subjects (mean ± sem). CD8 Treg markedly inhibited IgM, IgG, and IgA production at all B cell:CD8 Treg ratios.