Abstract: Two distinct murine cell-surface differentiation antigens, Mac-1 and LFA-1 (lymphocyte function-associated antigen 1), are compared here and shown to be related at the molecular level. Mac-1, defined by the M1/70 rat anti-mouse monoclonal antibody (MAb), is expressed on macrophages, natural killer cells and 50% of bone marrow cells, but not on B or T lymphocytes1–3. In contrast, the LFA-1 antigen, defined by the M7/14 rat anti-mouse MAb, is expressed on B and T lymphocytes and 75% of bone marrow cells, but not on thioglycollate-induced peritoneal exudate macrophages4,5. MAb blocking studies suggest that LFA-1 participates in T-lymphocyte-mediated killing and T-lymphocyte antigen-specific responses4,5. Mac-1 and LFA-1 have α-polypeptide chains of 170,000 and 180,000 molecular weight (Mr), respectively, and both contain β polypeptides of 95,000 Mr. This similarity prompted us to investigate their relationship. Mac-1 and LFA-1 have distinct cellular distributions, MAb-defined antigenic determinants and α-subunits, but have highly homologous or identical β-subunits as shown by tryptic peptide mapping. Moreover, they share some common antigenic determinants recognized by a polyclonal antiserum. Cross-linking studies show that in each antigen the subunits are noncovalently associated in α1β1 structures. Mac-1 and LFA-1 comprise a novel family of two-chain leukocyte differentiation antigens.

Abstract: The major histocompatibility complex (MHC)-linked immune response (Ir) genes confer high responsiveness on some MHC haplotypes and low or no responsiveness to certain antigens on other haplotypes (for review see ref. 1). The cause of nonresponsiveness is thought to be a failure of T cells to be stimulated by a given combination of antigen and MHC molecules, which can, in principle, arise in two ways: either the antigen-presenting cells (APCs or macrophages) of the nonresponder haplotypes are incapable of presenting the foreign antigen to T cells in an immunogenic form2,3 or the T-cell repertoire of nonresponder haplotypes lacks clones recognizing the antigen in the context of a given MHC4. We have recently shown that APCs from nonresponder strains can in fact present antigen to allogeneic responder T cells, depleted of alloreactivity against the APCs themselves5, which indicates that the mechanism of nonresponsiveness is not a failure of APC function. Using the same experimental system, we show here that responses of T cells to antigen presented on allogeneic APCs do not depend on the Ir phenotype of either the T cell or the APC: Ir gene control is completely absent from these responses. The results strongly suggest that Ir gene-controlled nonresponsiveness is associated with the adjustment of the T-cell repertoire to self-MHC antigens.

Abstract: We have investigated the ability of an Ia-, nonantigen-presenting macrophage tumor cell line, P388D, (H-2d), to present antigen to T cell hybridomas after incubation in a lymphokine-containing preparation. P388D, cells were incubated in microtiter wells with various concentrations of Con A-stimulated spleen cell supernatants. Antigen-specific stimulation of H-2d-restricted, KLH-specific T cell hybridomas was observed by P388D1 incubated with SUP.P388D1 cells incubated for 3 days in medium or control SUP did not present antigen. In addition, no stimulation of T hybridomas was seen by P388D1 in the inhibited by the appropriate monoclonal anti-Ia reagents. These results demonstrate that a macrophage tumor cell line can be induced to present antigen and provides for large numbers of readily available, homogeneous macrophages for studying the cellular biochemical requirements for antigen processing and presentation.

Abstract: The mechanisms responsible for the induction of I-J restrictions on third-order suppressor T cells (TS3) were analyzed. The I-J phenotype of the antigen-coupled cells used for priming restricted the specificity of the TS3 population. Thus, TS3 cells were only generated after priming with antigen-coupled I-J homologous cells. Identity at the I-JM (and I-E) subregions was sufficient for TS3 induction. Furthermore, priming of H-2 heterozygous mice with antigen-coupled parental cells generated TS3 that were restricted to the parental haplotype used for priming. The splenic cell population responsible for antigen presentation and induction of TS3 cells was fractionated. The cells involved in antigen presentation were found in the splenic adherent population and were absent in the fraction containing splenic nonadherent T and B cells. The subsequent activation and interaction of TS3 cells is also restricted by genes in the H-2 complex. The results are discussed in terms of a general mechanism responsible for the induction of restrictions in T helper and TS3 cells.

Abstract: We investigated the antigenic requirements for restimulation of H-2- restricted cytolytic T lymphocytes (CTL) in vitro to determine whether H-2 I region-restricted helper T cells are required in these responses. In one set of experiments, we studied the in vitro response of (responder x nonresponder)F(1) female T cells to the male antigen H-Y. We chose to examine this response because it has been suggested that the defect in nonresponder strains is a failure of helper T cells to recognize H-Y in association with nonresponder I region determinants. However, we find that nonresponder male stimulator cells are as effective as F(1) male stimulator cells at inducing H-Y-specific CTL responses. This finding calls into question reports that secondary CTL responses to H-Y are dependent upon the activation of H-Y- specific helper T cells restricted to responder type I region determinants. In a second set of experiments, we examined the requirements for restimulation of H-2-restricted T cells specific for minor-histocompatibility antigens from long-term mixed lymphocyte cultures. These cultures were established by repeatedly restimulating cultures of specific T cells with H- 2-matched stimulator cells expressing foreign minor histocompatibility antigens. We found that H-2D-restricted T ceils, including CTL, could be restimulated with cells that were matched with the responding cells at only the D region genes. This response did not appear to result from positive allogeneic effects or from antigen processing and “representation” by responder type APC that might contaminate the cultures. Thus, we find no evidence for a requirement for I region-restricted helper T cells in these CTL responses. However, helper T cells are required because we find that CTL lines derived by limit-dilution cloning from these long-term MLC are absolutely dependent upon exogenous helper factors for growth. The most simple interpretation of these results is that the helper cells are restricted to H-2 antigens other than I region antigens or to antigens that code outside of the H-2 complex. Finally, we show that factor-dependent CTL lines must recognize their specific antigen to proliferate, even in the presence of exogenous factors. The requirement of activated CTL for antigen to proliferate provides an explanation for how specific CTL can be selectively enriched in MLC by specific antigen stimulation. Furthermore, it is at variance with reports that memory CTL or activated CTL require only interleukin 2 for restimulation.