Interaction Between Lymphocytes and Inflammatory Exudate Cells. II. A Proteolytic Enzyme Released by PMN As a Possible Mediator for Enhancement of Thymocyte Response

Mouse polymorphonuclear leukocytes (PMN) were harvested from the peritoneal cavity stimulated with sodium caseinate. The cells were cultivated in vitro and the supernatant of these cultures (SUP) was tested for enhancing potency on DNA synthesis by syngeneic thymocytes stimulated with phytohemagglutinin (PHA). The enhancing potency of the SUP was markedly influenced by duration of the donor cultures of PMN, population density of the cultures, and protein concentrations in the medium, respectively. The enhancing factor in the SUP was found to be non-dialysable, heat-labile, and stable in the pH ranging between 3 and 9 but labile in the pH below 2 or above 10; its m.w. was approximately 19.000 when measured by gel-filtration on Sephadex G-75. The factor had a proteolytic activity on 3H-acetyl hemoglobin (3HHb) at neutral pH (7.2). Both the proteolytic activity and thymocyte-helping potency of the SUP were similarly abolished by adding protease inhibitor (Trasylol) in soluble form, or by passing through a column of the inhibitor insolubilized. It was thus assumed that the enhancing effect of PMN on thymocyte response was associated with a neutral protease released from the cells: it may be termed a lymphocyte-helping protease (LHP).