Regulation of lymphocyte proliferation by eosinophils via chymotrypsin-like protease activity and adhesion molecule interaction

1 We investigated the regulatory mechanisms responsible for release of eosinophil cationic protein (ECP) from eosinophils activated by platelet-activating factor (PAF) and monitored intra-cellular pH (pHi) changes using a pH-sensitive fluorescent probe. We also explored the mechanisms by which eosinophils suppress T-lymphocyte proliferation induced by phytohaemagglutinin (PHA). In these experiments, a separated culture to investigate the ECP-mediated pathway and a coculture to identify the adhesion molecules involved in eosinophil-lymphocyte interactions were employed. 2 Chymostatin (1 x 10(-6) M) inhibited ECP release by about 50% via stimulation by PAF or recombinant interleukin 5(rIL-5) plus IgG. 3 PAF (1 x 10(-7) M) raised eosinophil pHi from 6.9 to 7.3 within 20 a and pretreatment of these cells with chymostatin (1 x 10(-6) M), but not with leupeptin or E64-d, completely prevented this increase. 4 Calcium ionophore A23187 (1 x 10(-7) M) induced ECP release and raised pHi to within a range similar to that of PAF, however, chymostatin had no effect on either. 5 Chymostatin reversed ECP-mediated suppression of PHA-induced T-lymphocyte proliferation in separated cultures, but not in cocultures. 6 In coculture, eosinophils exhibited the same level of suppression of both CD4+ and CD8(+) T-cell proliferation in response to PHA. 7 Monoclonal antibodies against CD11a, CD18 and CD54, but not CD11b, restored eosinophil suppression of T-lymphocyte proliferation which was chymostatin-resistant in coculture. 8 Eosinophils were unable to suppress the proliferative response to lymphocytes to anti-CD3 stimulation. 9 In conclusion, chymostatin specifically inhibited both the eosinophil pHi increase and ECP release induced by PAF. Eosinophils regulate PKA-induced T-lymphocyte proliferation via the ECP-mediation associated with chymotrypsin-like protease activity. These cells also control interactions with lymphocyte between adhesion molecules, CD11a, CD18 and CD54.