A fast-acting cytotoxin derived from Con A-activated porcine leukocytes: II. Mechanism of target cell lysis

Abstract

Serum-free culture supernatants of Con A-stimulated pig leukocytes contain cytotoxins (PCT) which have a fast-acting lytic effect on a variety of murine lymphomas. Mathematical analysis revealed that the cytodestructive reaction which leads to the immediate release of 51Cr from labeled target cells follows “single hit”/“first order” kinetics. The release of labeled compounds was found to be size dependent, such that low molecular isotope ( 86Rb) was released at faster rates than high molecular 51Cr-labeled complexes. The 51Cr release was strongly reduced by lowering the temperature to 4 °C. PCT-mediated target cell lysis could be competed by cold target cells and the factor could be absorbed by intact cells, plasma membranes and artificial liposomes. Neither the presence of EDTA or of various monosaccharides, nor exposure of target cells to trypsin, metabolic inhibitors, and cytoskeletal antagonists altered the target cells susceptibility to PCT-mediated lysis. Labeling of target cell DNA and subsequent exposure of these target cells to PCT revealed that target cell DNA is degraded into low-molecular-weight split products which is similar to that seen during T-cell mediated target cell lysis. The analysis of the lytic event mediated by PCT thus revealed similarities to T-cell mediated target cell lysis. PCT was clearly distinct from the well-known cytolytic agents lymphotoxin (LT), tumor necrosis factor (TNF), or complement (C′). Taken together, the described characteristics make PCT a unique-acting cytolytic cytokine with anticancer activity and suggest further research concerning its mode of action and its possible therapeutic application.