ENHANCEMENT OF PHYTOHEMAGGLUTININ‐INDUCED DNA SYNTHESIS OF MOUSE THYMOCYTES AND T‐LYMPHOCYTES BY A NEUTRAL PROTEASE OF POLYMORPHONUCLEAR LEUKOCYTES

Abstract

Mouse thymocytes were poorly triggered in vitro with phytohemagglutinin (PHA) to induce DNA synthesis and the response was markedly enhanced by culture supernatant (SUP) of polymorphonuclear leukocytes (PMN). The PMN factor was nondialysable, heat-labile and precipitated with 65% ammonium sulphate. Its molecular weight was approximately 19,000 on sephadex G-75. The sephadex fraction had a proteolytic activity on 3H-acetyl hemoglobin at neutral pH. The protease seemed to be a chymotrypsin-like enzyme on the basis of inhibition profile using various protease inhibitors. The thymocyte-helping activity in the protease fraction was absorbed by affinity columns of protease inhibitors. The PMN-protease also enhanced the DNA synthesis by antigen-stimulated lymph node cells and by PHA-stimulated T-lymphocytes, but not by LPS-stimulated B-lymphocytes. Only the lymphocytes already been stimulated with antigen or mitogen received preferentially the helping action by PMN-protease. The helping activity was effectively absorbed by PHA-stimulated thymocytes but not by nonstimulated thymocytes. These evidences seems to suggest that acceptor sites for the protease, newly developed on lymphocyte surface after stimulation, may play an important role in the enhanced DNA synthetic response.