A comparison of lysis mediated by Lyt 2+ TNP-specific cytotoxic-T-lymphocyte (CTL) lines with that mediated by rapidly internalized lymphotoxin-containing supernatant fluids: evidence for a role of soluble mediators in CTL-mediated killing.

A series of Lyt 2+, trinitrophenyl (TNP)-specific T-cell lines are shown to lyse 51Cr-labeled target cells in an antigen-specific, H-2-restricted fashion in a 4-hr assay. These cells also produce lymphotoxin, in addition to other factors, upon stimulation with TNP-haptenated syngeneic splenocytes. A technique for introducing macromolecules into the cytoplasm of fibroblasts by inducing the cells to pinocytose the molecule in hypertonic medium, and then lysing the newly formed pinocytic vesicles with a mild hypotonic shock was used to assess the role of soluble mediators in the cytotoxic T lymphocyte (CTL)-mediated lytic process. The technique itself has little effect on cell growth rate or viability. A minimum of 24 hr, and more frequently 48-72 hr is required for lymphotoxin to manifest it's lethal effect when it is merely included in the culture medium of growing fibroblasts. In contrast, supernatant fluids from the Lyt 2+ cells kill 51Cr-labeled fibroblasts in a dose-dependent fashion during a 4-hr assay when they are rapidly internalized via the osmotic procedure. The data serve as preliminary evidence of a role for soluble mediators such as lymphotoxin in T-cell-mediated lysis, and suggest that the cytotoxic-T-cell lethal hit may include a mechanism for rapidly internalizing a toxin into appropriate target cells.