Inhibition of Stimulation in Murine Mixed Lymphocyte Cultures with an Alloantiserum Directed Against a Shared Ia Determinant

Abstract

Pools of high titered alloantisera were raised by immunizing (B10.A/SgSn × A/WySn)F₁ mice with C57BL/10Sn(B10) spleen cells. This serum (F₁ anti-B10), when added to one-way mixed lymphocyte cultures (MLC), inhibited stimulation of B10.A splenic responders by both B10 and B10.D2/nSn irradiated, splenic stimulators. The B10 stimulation was suppressed approximately 85% whereas the mean suppression of B10.D2 stimulation was approximately 60%. In the former case, the serum contained antibodies reactive with multiple major histocompatibility complex determinants on the stimulator cells. In the latter case, the cytotoxic reactivity of the serum was directed principally against an I region-associated determinant (Ia.8) shared by B10 and B10.D2 and coded for by a gene(s) in the I-A subregion. The magnitude of the suppression of the response to B10.D2 cells (60%) was similar to the reduction in stimulation observed when the Ia.8 difference was eliminated genetically by using (B10 × B10.A)F₁ responder cells against irradiated B10.D2 stimulators. Inhibition of MLC by this antiserum was a function of reactivity with stimulator and not responder cells. Although some pools of F₁ anti-B10 antiserum produced partial inhibition of the responder cell in a B10.D2 vs B10.A_x MLC combination, the results were inconsistent and not correlated with the anti-Ia.8 cytotoxicity titers. In addition, an F₁ anti-B10 antiserum pool, which consistently failed to inhibit the responder cell, nevertheless inhibited both irradiated B10.D2 and (B10.A × B10.D2)F₁ cells from stimulating B10.A responder cells. However, this same antiserum did not inhibit stimulation of B10.D2 responder cells by the (B10.A × B10.D2)F₁ stimulators. Thus, the binding of antibodies to the non-stimulating antigens on the F₁ stimulator cell did not interfere with the capacity of the appropriate stimulating antigens to cause stimulation. All of these results are consistent with the hypothesis that Ia alloantigens are the major stimulating determinants of I region-associated MLC reactions.