Alloantigen-induced T-cell proliferation: Lyt phenotype of responding cells and blocking of proliferation by Lyt antisera

Cytotoxic T cells of the mouse express Lyt-1 as well as Lyt-2 and -3 on their surface, and T-cell cytotoxicity can be blocked by Lyt-2 and Lyt-3 (but not Lyt-1) antisera in the absence of added complement [Nakayama, E., Shiku, H., Stockert, E., Oettgen, H. F. & Old, L. J. (1979) Proc. Natl. Acad. Sci. USA 76, 1977-1981]. This analysis has now been extended to the study of the Lyt phenotype of T cells responding to alloantigens, concanavalin A (Con A), and phytohemagglutinin (PHA) and the effect of Lyt antibody on T-cell proliferation and the generation of H-2-specific killer T cells. H-2 (D/K and I), Con A, and PHA stimulation was abolished by pretreating responding cell populations with Lyt-1 antiserum and complement. Pretreatment with Lyt-2 or -3 antiserum and complement did not decrease alloantigen or Con A stimulation but did abolish PHA stimulation. Cytotoxic cells were not generated in H-2 alloantigen-primed cultures pretreated with Lyt-1, -2, or -3 antiserum and complement. When responding cells were cultured with Lyt antiserum in the absence of added complement, Lyt-2 or -3 antiserum (but not Lyt-1 antiserum) blocked alloantigen-induced proliferation and delayed generation of killer cells. Under similar conditions, Con A and PHA stimulation was not blocked by Lyt-1,-2, or -3 antiserum. Evidence from these Lyt elimination and blocking tests and from direct Lyt phenotyping of responding cells leads to the following conclusions. Two populations of Lyt(+) cells are involved: Lyt-1(+)2(-)3(-) and Lyt-1(+)2(+)3(+). Current evidence does not favor the existence of Lyt-1(-)2(+)3(+) cells but indicates that pre-killer and killer cells derive from the Lyt-1(+)2(+)3(+) population and have a Lyt-1(+)2(+)3(+) phenotype. H-2 (D/K and I) and PHA stimulation ordinarily activate the Lyt-1(+)2(+)3(+) population, whereas Con A and I region or Mls locus antigens activate the Lyt-1(+)2(-)3(-) population. However, when Lyt-1(+)2(+)3(+) cells are eliminated or blocked by Lyt-2 or -3 antiserum, H-2 alloantigen stimulation leads to proliferation of the Lyt-1(+)2(-)3(-) population. Blocking of H-2-induced proliferation by Lyt-2 or -3 antiserum adds further support to the possibility that molecules bearing Lyt-2 and -3 determinants are involved in T-cell recognition

Cytotoxic T cells: Lyt phenotype and blocking of killing activity by Lyt antisera

We reexamined two questions concerning Lyt antigens of cytotoxic T cells of the mouse: is Lyt-1 antigen expressed on cytotoxic effector cells and can cytotoxicity be blocked by antibody to Lyt antigens in the absence of added complement? A 3-hr (51)Cr-release assay with splenic effector cells and leukemia or myeloma target cells was used to measure cell-mediated cytotoxicity. The cytotoxic activity of effector cells against allogeneic targets was abolished by exposure to Lyt-1, Lyt-2, or Lyt-3 antiserum and complement. Specificity was established by tests with C57BL/6 Lyt congenic mice and absorption studies with thymocytes. Similarly, the cytotoxicity of effector cells directed against semisyngeneic myeloma targets was reduced by Lyt-1, -2, or -3 antiserum and complement. Effector cell cytotoxicity against another semisyngeneic target was only marginally affected by Lyt-1 antiserum and complement, but was abolished by Lyt-2 or -3 antiserum and complement. It appears likely that cytotoxic T cells are a heterogeneous population with regard to Lyt-1 expression and that past studies indicating an apparent absence of Lyt-1 on cytotoxic T cells revealed a quantitative, not qualitative, feature of these cells. With regard to the activity of Lyt antisera in the absence of added complement, selective blocking of effector cell cytotoxicity for allogeneic and semisyngeneic targets was found with Lyt-2 and Lyt-3 antisera but not with Lyt-1 antiserum. The specificity of blocking was established by tests with Lyt congenic mice and absorption studies with thymocytes. With the exception of blocking by antisera to the H-2 haplotype expressed by the target cell, no effector cell blocking was observed with alloantisera or heteroantisera to a range of other cell surface molecules present on mouse lymphoid cells. One possibility to account for the selective blocking by Lyt-2 and Lyt-3 antisera is that Lyt-2,3 determinants on the surface of cytotoxic T cells have a close spatial relation to the T cell receptor