LYSIS OF ENUCLEATED TUMOR-CELLS WITH ALLOGENEIC AND SYNGENEIC CYTOTOXIC THYMUS-DERIVED LYMPHOCYTES

These studies were initiated to understand the minimal requirements for lysis of target cells by cytotoxic thymus‐derived lymphocytes (CTL). In particular, the significance of the nucleus to the susceptibility of the target cell to be lysed by CTL have been studied. P815 mastocytoma cells and EL4 lymphoma cells were enucleated by cen‐ trifugation through a discontinuous Ficoll gradient containing 10 μg/ml of cytochalasin B. The resultant vesicles were then analyzed for susceptibility to lysis by different CTL specific for the H‐2K and H‐2D gene products, minor histocompatibility antigens, trinitrophenyl groups, and Sendai virus‐specific antigens. The results indicate that enucleated vesicles obtained from both EL 4 and P 815 cells are susceptible to lysis by these CTL. The lysis of EL 4‐enucleated vesicles (unlike P 815‐enucleated vesicles), however, required 2–4 times more effector cells than were required for equivalent lysis of intact EL 4 cells. This reduced susceptibility to lysis of enucleated EL 4 vesicles was not the result of an inability of the CTL to recognize and bind the vesicles. This conclusion was suggested by the fact that both enucleated EL 4 vesicles and intact cells inhibited equally well the lysis of 51Cr‐labeled concanavalin A‐stimulated BALB. B spleen cells by anti‐H‐2b CTL. Although enucleated vesicles obtained from both P 815 and EL 4 cells were susceptible to lysis by CTL, they exhibited a reduced capacity to “cap” their serologically defined H‐2 antigens. Lysis of these enucleated vesicles by CTL has the same requirements as lysis of intact cells. This conclusion was based upon the requirement for Ca++ and Mg++ and also because lysis of the modified vesicles (trinitrophenyl or viral antigens) occurred in an H‐2‐restricted fashion. Moreover, the enucleated vesicles were susceptible to antibody and complement‐mediated lysis and also lectin‐dependent cell‐mediated cytotoxicity. Copyright © 1979 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim