Cell‐free media of mixed lymphocyte cultures augmenting sensitization in vitro of mouse T lymphocytes against allogeneic fibroblasts

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Abstract
Using an in vitro mouse lymphocyte anti‐fibroblast reaction (AFR), we recently reported that the addition of allogeneic stimulator lymphocytes to the sensitization phase of the AFR enhanced sensitization to fibroblast antigens as evidenced by a marked increase in the cytolytic activity of the sensitized lymphocyte population. In the present report we studied the mechanism of this helper effect by testing the capacity of cell‐free media derived from 48‐h mixed spleen cultures to enhance anti‐fibroblast sensitization. We found that such cell‐free media could produce a marked helper effect when applied to the sensitization phase of the AFR, but not when added to the cytolytic effector phase. The stimulator cells in the mixed lymphocyte culture (MLC) did not have to be syngeneic with the sensitizing fibroblasts in the AFR in order for the helper effect to be demonstrated. Lymphocytes sensitized to fibroblast antigens in the presence of MLC medium retained their specificity of the effector phase. Our data suggest that the MLC medium acts by enhancing differentiation processes of antigen‐triggered lymphocytes. Generation of helper activity by the MLC was abolished by 1000 r irradiation of the responder cells. By using nu/nu and normal spleens, both from two different strains, as cell sources for the MLC, we found that the generation of helper activity depended on T cells capable of proliferation. Furthermore, stimulator lymphocytes differing from responder lymphocytes by non‐H‐2 alloantigens as well as by point mutation within the H‐2 complex were capable of eliciting helper factor(s). Thus, soluble factor(s) produced in a MLC, which are dependent on T lymphocyte proliferation, have the capacity to enhance the sensitization of mouse lymphocytes against antigens present on allogeneic fibroblasts.