ROLE OF FUNCTION-ASSOCIATED MOLECULES IN TARGET-CELL LYSIS - ANALYSIS OF RAT ADHERENT LYMPHOKINE-ACTIVATED KILLER-CELLS

Monoclonal antibodies (MAbs) against fish nonspecific cytotoxic cells (NCC) inhibit the lysis of target cells by NCC and by human and rat natural killer (NK) cells. We now show that the anti-function-associated molecule (anti-FAM) MAb 5C6 also inhibits the cytolytic activity of rat (F344) nonadherent lymphokine-activated killer (LAK) cells and adherent LAK (ALAK) cells. Rat nylon-wool-nonadherent (NWNA) cells were cultured for 48 h in recombinant human interleukin-2 (IL-2). Nonadherent cells were then removed and the plastic-adherent (ALAK) cells were recultured for 9 days. Cells were daily analyzed by cytometry (using MAb 5C6 and the rat NK cell MAb 3.2.3) and cytotoxicity. Approximately 40% of the initially cultured NWNA cells, 50-60% of the LAK cells and 87-98% of the ALAK cells were positive for MAb 5C6. Tissue culture supernatants containing MAb 5C6 caused a dose-dependent inhibition of cytotoxicity against P815 and YAC-1 targets. Approximately 50% inhibition was obtained at MAb concentrations of 1.25-2.5 mug/ml. Maximum inhibition of ALAK cytotoxicity was 86% on day 4 of culture. Two-dimensional SDS-PAGE and Western blot analysis of acetone-precipitated cell membrane preparations identified a 55-kD molecule (pI 6.1-6.3) on NWNA and ALAK cells and an additional slightly more acidic 45-kD protein on ALAK cells. These data indicate that rat ALAK cells may initiate lysis of target cells via receptor binding.