Conditions for the enhancing effect of protease inhibitors on the concanavalin A induced thymidine response of murine lymphocytes.

Abstract

Incorporation of [3H]-thymidine - [3H]-TdR - into concanavalin A (Con A) stimulated murine splenocytes and thymocytes was found to be enhanced by addition of certain concentrations of phenyl-methylsulfonylfluoride (PMSF), di-isopropylfluorophosphate (DFP), N-alpha-tosyl-L-lysyl-L-chloromethylketone (TLCK), and soybean trypsin inhibitor (SBTI). No enhancement could be observed when mononuclear cells of the peripheral blood were used, and a medium enhancement when thymocytes were applied. Furthermore, no enhancing effect of the protease inhibitors (PI) on the Con A response of murine splenocytes could be observed within the first 24 h of the culturing period. DFP, PMSF, and TLCK enhanced the Con A response to a similar degree, whereas SBTI was less effective. DFP and SBTI proved to be also effective when they were added after 15-24 h to the Con A cultures, if the cultures were harvested 48 h later. Removal of adherent and phagocytic spleen cells or reduction of the concentration of spleen cells shifted the effective DFP concentration to lower concentrations, whereas addition of adherent spleen cells caused a shift of the enhancing DFP amounts to higher concentrations. The data presented suggest that the enhancing effect of PI on the T cell response depends on the concentration of PI, the time of culturing and incubation, the PI used, the origin of the stimulated cells, and especially on the number of adherent and phagocytic cells. These findings might explain - at least in part - the different results on the effect of PI on the T cell response obtained in the past.