In vitro determination of self-reactivity in the early postcyclosporine period

Abstract

Since cyclosporine A(CsA) was introduced into transplantation medicine to prevent graft-versus-host disease (GvHD) as well as graft rejection, side-effects became obvious. When CsA is given and withdrawn GvHD-like symptoms can occur even in an autologous setting. To understand this mechanism we tested the allo- and self-reactivity of murine spleen lymphocytes in an in vitro assay. Mice of four different strains with two distinct MHC class I backgrounds (H-2 d and H-2 k) were treated with 60 mg/kg/day CsA intraperiteonally for ten days. In an attempt to examine the possibility that the CsA-induced autoimmunity requires the presence of the thymus, half of these four- to six-week-old mice were also thymectomized prior to the CsA treatment. Within one day after CsA was stopped, all mice that received CsA during treatment showed reactivity against self-MHC-bearing spleen cells. This was demonstrated in a primary in vitro stimulation assay followed by a chromium-release assay (H-2 d-anti-H-2 d and H-2 k-anti-H-2 k). However, alloreactivity (H-2 d-anti-H-2 k and H-2 k-anti-H-2 d) was suppressed. At this point in time, no natural killer (NK) activity was detectable in any of the CsA-treated mice. The days after stopping CsA, the autoreactivity was no longer detectable in any mouse strain, whether thymectomized or not, whereas the alloreactivity and the NK activity finally recovered. The in vitro phenomena of self-reactivity, which occurred between day one and day 10 after CsA withdrawal, could be adoptively transferred from syngeneic in vitro reactive T cell populations into H-2 identical mice. The sublethally irradiated recipients of autoreactive cells died within 10–20 days, showing clinical signs of pseudo GvHD (pGvHD). Irradiated control mice not receiving self-reactive T cells, and mice that received the self-reactive T cells but were not irradiated, survived without complications. Our results suggest the existence of self-reactive MHC-restricted T cells which, under normal immunological conditions, are regulated and probably nonfunctional in vivo. However, CsA treatment or other immunomodulatory manipulations may interfere with the maintainance of self-tolerance and lead to the activation of these self-reactive T cells. The role of the thymus at this stage of T cell selection during CsA treatment is not clear, since we could show that CsA has also an effect on mature T cells in the periphery.