In B-cell chronic lymphocytic leukemia (B-CLL) microenvironments, interplaying between tumor necrosis factor (TNF) superfamily members and their cognate receptors has been shown to play a relevant role in controlling B-CLL growth and survival. Death Receptor 3 (DR3), a TNF receptor superfamily member, has been recently associated with modulation of immune and inflammatory functions. The ligand of DR3, TNF-like protein 1A (TL1A), is expressed by various cell types, including bystander cells in B-CLL microenvironments, i.e. macrophages and activated T cells. The aim of this study was to test the hypothesis that DR3 may be involved in the molecular network that modulates leukemic growth in B-CLL microenvironments. The expression of DR3 was measured in peripheral blood B cells and in lymph node specimens from 35 B-CLL patients by flow cytometry and immunofluorescence, respectively. PBMCs and lymph nodes from healthy donors were used as control. Although B cells from either B-CLL patients or healthy donors did not express DR3 in basal conditions, stimulation of B cell receptor (BCR) with anti-IgM antibodies induced de novo expression of DR3 in B cells from both B-CLL patients and healthy donors. However, DR3 expression was significantly higher in B cells from B-CLL patients than that in healthy donors (mean fold-induction ± SD = 2.51±1.1 in B-CLL and 1.91±0.5 in healthy donors; p 0.05). DR3 expression in B-CLL cells was confirmed at the mRNA level by quantitative RT-PCR (4-fold induction with respect to basal conditions). Immunofluorescence analysis showed DR3 expression both in scattered CD20 positive cells in B-CLL lymph node and in CD20 positive cells in germinal centers of reactive lymph nodes. DR3 functional activity was analyzed in anti-IgM-stimulated BCLL cells by engaging DR3 with agonistic antibodies. DR3 engagement induced signaling activity, as shown by the downstream phosphorylation of Extracellular Signal-Regulated Kinase 1 and 2 (Erk1/2) measured by phospho-specific flow cytometry. In conclusion, we described for the first time the expression of functional DR3 molecules in B-CLL cells. The finding that anti-IgM-induced DR3 expression was higher in B-CLL cells if compared with healthy B cells suggests that DR3 may have significance in B-CLL physiopathology.
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