CD70+ non-Hodgkin lymphoma B cells induce Foxp3 expression and regulatory function in intratumoral CD4+CD25 T cells - PubMed (original) (raw)

CD70+ non-Hodgkin lymphoma B cells induce Foxp3 expression and regulatory function in intratumoral CD4+CD25 T cells

Zhi-Zhang Yang et al. Blood. 2007.

Abstract

Foxp3 expression was initially thought to be restricted to the CD4(+)CD25(+) regulatory T-cell population. However, recent studies suggest that forkhead box P3 (Foxp3) is expressed in CD4(+)CD25(-) T cells in aged mice. In the present study in B-cell non-Hodgkin lymphoma (NHL), we found that a subset of intratumoral but not peripheral blood CD4(+)CD25(-) T cells, comprising about 15% of intratumoral CD4(+) T cells, express Foxp3 and are capable of suppressing the proliferation of autologous infiltrating CD8(+) T cells. In vitro activation with OKT3/anti-CD28 antibody (Ab) or dendritic cells (DCs) induced Foxp3 expression in a subset of these CD4(+)CD25(-)Foxp3(-) T cells. We found that the presence of lymphoma B cells during activation augmented activation-induced Foxp3 expression in CD4(+)CD25(-) T cells. We also found that CD70(+) lymphoma B cells significantly contributed to the activation-induced Foxp3 expression in intratumoral CD4(+)CD25(-) T cells. Furthermore, the blockade of CD27-CD70 interaction by anti-CD70 Ab abrogated lymphoma B-cell-mediated induction of Foxp3 expression in intratumoral CD4(+)CD25(-) T cells. Taken together, these studies reveal a novel role for NHL B cells in the development of intratumoral regulatory T cells.

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Figures

Figure 1

Foxp3 expression in CD4+CD25− T cells in biopsy specimens of B-cell NHL. (A) Representative dot plots showing Foxp3 expression in CD4+CD25+ and CD4+CD25− T cells in biopsy specimens of patients with NHL. Intracellular Foxp3 expression was determined by using fluorochrome-conjugated Foxp3 antibody. (B) Percentage of Foxp3 expression in different subsets of cells in NHL specimens (n = 12). The horizontal bars indicate median expression levels. (C) Representative dot plots showing Foxp3 expression in positive (CD25+) and negative (CD25−) fractions of CD25-depleted sample. (D) Representative immunofluorescence images showing Foxp3 expression in CD25+ and CD25− cells in biopsy specimens of patients with NHL. Tissues were stained with antibodies to human CD25 and Foxp3. Left panels shows isotype control for each antibody; right, double immunofluorescence staining of CD25 (red; surface) and Foxp3 (green; intracellular). Original magnification, × 200. See “Immunofluorescence assay” for image acquisition details. (E) Representative dot plots showing Foxp3 expression in CD4+CD25− T cells (CD25-depleted) from tumor tissue (top panel) and peripheral blood mononuclear cells (bottom panel) from patients with NHL. Percentages in panels A,C, and E represent number of cells in quadrant as a percentage of all cells.

Figure 2

Regulatory function of CD4+CD25− T cells in biopsy specimens of B-cell NHL. CFSE-labeled CD8+ T cells were cocultured either alone or with intratumoral CD4+CD25− T cells or intratumoral Treg cells (CD4+CD25+) in the presence (Stim) or absence (Unstim) of PHA for 3 days. Proliferation of CD8+ T cells is measured based on CFSEdim cells. The figure shown is representative of 3 independent experiments (3 samples) with similar results. Percentages indicate number of proliferated CD8+ T cells (CFSEdim) of just the CD8+ cell population.

Figure 3

Foxp3 induction in CD4+CD25− T cells in biopsy specimens of B-cell NHL. Foxp3 expression was examined in CD4+CD25− T cells stimulated with OKT3 or OKT3/anti-CD28 Ab (A) (n = 10) or with in vitro–generated DCs pulsed with or without super-Ag TSST-1 (B) (n = 3). (C) Foxp3 expression in CD4+CD25− or CD8+ T cells upon activation. CFSE was used to monitor activation and proliferation (n = 6). Percentages indicate number of proliferated CD8+ T cells (CFSEdim) of just the CD8+ cell population. (D) CD25 and Foxp3 expression determined at specific day after activation in intratumoral CD4+CD25− T cells in B-cell NHL (n = 4).

Figure 4

Effect of lymphoma B cells on activation-induced Foxp3 expression in CD4+CD25− T-cell biopsy specimens of B-cell NHL. (A) Purified CD4+CD25−CD127+ T cells were cultured in OKT3-coated plates (OKT3) or with the addition of anti-CD28 Ab (OKT3/aCD28) or CD19+ lymphoma cells (OKT3 + LB) for 3 days. Unstimulated cells (Resting) were used as the baseline for calculating Foxp3 induction. Foxp3 expression was examined by intracellular staining. Numbers indicate CD4+Foxp3+ cells as a percentage of all cells. (B) Summarization of Foxp3 induction in CD4+CD25− T cells upon activation with OKT3 alone (OKT3; n = 15) or plus anti-CD28 (+ aCD28; n = 6) or in the presence of lymphoma B cells (+ LB; n = 15). Foxp3 induction is indicated by fold change to group without stimulation (Resting). The horizontal bars indicate median.

Figure 5

Effect of CD27-CD70 interaction on Foxp3 induction in CD4+CD25− T-cell biopsy specimens of B-cell NHL. (A) Representative dot plots showing CD70 expression on CD19+ B cells in biopsy specimens from patients with lymphoma or biopsy specimens from patients with follicular hyperplasia or PBMCs from healthy individuals. Percentages indicate number of CD19+CD70+ cells as a percentage of all unsorted mononuclear cells. (B) Summarization of frequency of CD19+CD70+ cells in biopsy specimens of patients with NHL (n = 11). The horizontal bar represents the median Foxp3 expression level. (C) CD70+ (CD70+ LB) and CD70− (CD70− LB) lymphoma B cells were isolated by flow sorting and cocultured with intratumoral CD4+CD25− T cells in plates coated with (OKT3) or without (R) OKT3 for 3 days. Foxp3 expression was determined by intracellular staining. (D) Summarization of Foxp3 induction in CD4+CD25− T cells cocultured with lymphoma B cells (LB) treated with anti-CD70 Ab alone (LB/anti-CD70), or combination with anti-CD80 and CD86 Abs (LB/anti-CD70 + CD80CD86) in plates precoated with OKT3 for 3 days. Foxp3 expression was determined by intracellular staining. Foxp3 induction is indicated by fold change to group without stimulation (mean ± SE). *P < .05 compared with group with LB alone; **P < .01 compared with group with LB alone.

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CD70+ non-Hodgkin lymphoma B cells induce Foxp3 expression and regulatory function in intratumoral CD4+CD25 T cells - PubMed (original) (raw)