WHAT IS THE NATURE OF THE ANTIGENIC COMPLEX RECOGNIZED BY T LYMPHOCYTES?

Abstract

ABSTRACT: Macrophages modified by the trinitrophenyl (TNP) hapten have been used to clarify the nature of the antigenic determinant recognized by the antigen specific guinea pig T lymphocyte. We have previously demonstrated that the genetic restriction on the interaction of the TNP-modified macrophage and primed T lymphocyte was regulated by the Ia antigens of the macrophage used for initial sensitization and that following removal of alloreactive cells, T cells could be sensitized to TNP-modified allogeneic macrophages. In the present report, we will review our recent studies which have been directed to a further dissection of the immunogenic complex recognized by the antigen-specific T cell. It was relatively easy to define the role of macrophage Ia antigens in the complex as the antigen specific response to TNP-modified allogeneic macrophages was eliminated by anti-Ia sera directed against the allogeneic stimulator macrophage, while anti-Ia sera directed solely against the responder T cell produced no inhibition of the proliferative response. Treatment of TNP-modified macrophages with anti-Ia serum either immediately after or 24 hours after TNP modification resulted in a markedly deficient antigen-presenting cell. As brief treatment of the TNP-modified macrophage with anti-Ia serum did not inhibit Ia antigen synthesis, it is possible that treatment with anti-Ia serum resulted in removal of TNP determinants which had become associated with Ia antigens on the macrophage surface. It has been somewhat more difficult to clarify the contribution of antigenic determinants to the immunogenic complex recognized by the T cell. A marked reduction in the stimulatory capacity of freshly modified cells could be achieved by brief treatment with anti-TNP serum, but the stimulatory capacity of macrophages which had been TNP-modified and then “aged” for 24 hours prior to addition to primed T cells was unaffected by similar treatment. This result initially suggested that macrophage presentation of the TNP determinant is not simply a surface display phenomenon and that the macrophage must process membrane conjugated TNP in a manner so that it is inaccessible to anti-TNP antibody to create the relevant immunogen recognized by T cells. Recently, we have been able to demonstrate that anti-TNP antibody can inhibit the proliferative response induced by aged TNP-modified macrophages provided that they have been freshly modified with the non-cross reactive dinitrophenyl (DNP) hapten. Under these conditions sufficient antigen is available to allow binding of anti-TNP antibody and a highly specific inhibition of the proliferative response is observed. This experiment suggests that antigenic determinants are displayed on the macrophage surface, but under normal conditions at too low a density to allow efficient binding of anti-antigen antibody. In order to directly investigate the relationship between the TNP moiety and macrophage Ia antigens, macrophages were TNP-modified, radioiodinated, and lysed in detergent. When TNP-derivatized proteins were isolated using an anti-TNP immunoabsorbent and the presence of TNP-derivatized antigens in the eluted proteins determined by immunoprecipitation techniques, no hapten modified Ia antigens were detected. Furthermore, when Ia antigens from TNP-modified cells were eluted from an anti-Ia immunoabsorbent, no protein other than Ia antigens was detectable. Thus, although the functional studies with anti-Ia serum are strongly in favor of the model that TNP-conjugated membrane proteins are complexed with Ia antigens on the surface of the macrophage, we have not been able to prove the existence of this complex by the biochemical techniques employed. Nevertheless, a complex maintained by the integrity of an intact cell membrane might very well exist.