Eosinophil cdTCR Expression Figure 7. Specific mediator release upon CD3- or cdTCR-induced eosinophil activation. (A–C) Representative time course of ROS production following eosinophil activation by immunobilized specific antibodies (A), BrHPP (B) or SBA stimulation (C). (A) Stimulation with immobilized anti-CD3 (&), anti-cdTCR (X) or anti-Vd1 (m) mAbs or control IgG1 (¤). (B) Stimulation by BrHPP 200 nM (#) 100 nM (&) and 50 nM (e) or culture medium (¤). (C) Stimulation by SBA 500 mM (%) 50 mM (m) and 5 mM (X), culture medium (¤) or IgA/anti-IgA immune complexes (N). (D–F) EPO release by eosinophils upon CD3 or cdTCR activation by specific antibodies (D) (n = 6), BrHPP (E) (n = 3) or SBA stimulation (F) (n = 3). Results are expressed in count per second (cps). (G) EDN release by eosinophils upon CD3 or cdTCR activation by specific antibodies (n = 6). (H–I) Cytokine production by eosinophils upon CD3 or cdTCR activation by specific antibodies or IgA/anti-IgA immune complexes. IFN-c production (H) (n = 5–7) and TNF-a production (I) (n = 4–8). Results presented in panels E and F were obtained on different patients. *, P,0.05; **, P,0.01. doi:10.1371/journal.pone.0005926.g007 as well as the fragility of eosinophils was a constraint to obtain significant amounts of viable cdTCR+ eosinophils following cell sorting. Eosinophils display a rearranged cdTCR with a more restricted repertoire compared to cdT cells, further excluding the possibility of lymphocyte contaminants in eosinophil preparations. Besides T lymphocytes, dTCR rearrangements have only been reported in malignant B lymphoblasts [43], NK cells [44] and in mixed-type leukaemia retaining both T-lymphoid and myeloid characteristics. Furthermore, presence of double positive Vd1+Vd2+ and equal numbers of Vd1- and Vd2-expressing eosinophils in most of normal donors suggests incomplete allelic exclusion as found in some instances in cdT cells [45]. Direct cdTCR activation of eosinophils by cdTCR/CD3-specific antibodies and selective agonists induced eosinophil-specific responses. Release of such myeloid-associated mediators and the scarcity of circulating cd T cells further exclude the possibility that our results obtained using highly purified eosinophils would be due to cdTCR expression by contaminating lymphocytes. Thus, mature eosinophils expressing lymphoid markers might represent hitherto unattended cdTCR-expressing myeloid cells. Our results might be in agreement with observations that myeloid- derived plasmacytoid DC are able to express a ‘‘lymphoid program’’ (RAG1, rearranged IgH genes) [46]. By contrast, in the recently proposed myeloid-based model of haematopoiesis, T- or B-committed progenitors would each keep a myeloid- differentiation potential [47]. For instance, genetic reprogramming from differentiated B cells to macrophages has been reported [48]. Further experiments are required to unravel by which mechanisms eosinophils have acquired this lymphoid program. Of note, eosinophils are found in human thymus [8]. Furthermore, similarly to gut intraepithelial cdT cells, eosinophils are able to undergo peripheral differentiation, for instance in allergic lungs [49]. Investigating TCR expression by eosinophils in various PLoS ONE | www.plosone.org 10 June 2009 | Volume 4 | Issue 6 | e5926