CD28-MEDIATED CYTOTOXICITY OF YT NATURAL-KILLER-CELLS ON B7-POSITIVE TARGETS INDUCES RAPID NECROTIC DEATH INDEPENDENT OF GRANULE EXOCYTOSIS

The mechanisms leading to target cell killing by the human NK-like cel l line YT2C2 have been studied. YT2C2 cells express CD28 antigen and k ill B7-expressing targets by a CD28-mediated mechanism which is inhibi ted by anti-CD28 mAb (CD28.2). The lysis of B7-negative targets, which are also killed by YT2C2, is insensitive to CD28.2, but can be inhibi ted by cyclosporin A (CsA). CsA reduces degranulation in YT2C2 as meas ured by BLT-esterase release assays. A total suppression of B7-negativ e cell lysis was observed in the presence of EGTA, which blocks both d egranulation and perforin polymerization, confirming that lysis of thi s type of target depends solely upon granule exocytosis. In contrast, an additional extracellular EGTA-resistant component in B7-positive ta rget killing was evidenced. These results were consistently obtained w ith a panel of B7-positive and B7-negative targets, including a Jurkat subclone transfected to express B7 and its parental cell line. Ca2+-i ndependent killing was completed during the first hour of the cytotoxi city assay, whereas EGTA-sensitive lysis increased throughout the whol e incubation time. These two lytic mechanisms used by YT2C2 were found to induce two different modes of cell death. Extracellular Ca2+-depen dent killing caused apoptotic death in both B7-positive and B7-negativ e targets, whereas the EGTA-resistant cytolytic pathway, observed excl usively with B7-positive targets, led to necrosis. CD28 triggering in YT2C2 induces, therefore, an additional mechanism of cell killing, ind ependent of granule exocytosis, the nature of which remains to be iden tified. (C) 1996 Academic Press, Inc.