Isolation from Trypanosoma cruzi-infected mice of CD8+, MHC-restricted cytotoxic T cells that lyse parasite-infected target cells

Abstract

Recent in vivo depletion studies in mice demonstrated that CD8+ T cells play a critical role in acute resistance to experimental Trypanosoma cruzi infections. As part of efforts to characterize these protective CD8+ T cell effector populations, we report here that splenic lymphocytes from mice chronically infected with T. cruzi can be induced to express high levels of cytolytic activity after stimulation in vitro with irradiated T. cruzi-infected macrophages. Cytolytic activity can either be detected using a nonspecific lectin-dependent 51Cr-release assay or using 51Cr-labeled T. cruzi-infected target cells. Fresh splenocytes from chronically infected mice stimulated with T. cruzi-infected macrophages exhibit relatively "promiscuous" killing activity inasmuch as significant lysis of both T. cruzi-infected and uninfected syngeneic and allogeneic cells is detected. However, subsequent rounds of in vitro stimulation lead to the expression of lytic activity that is T. cruzi Ag-specific and MHC-restricted. Several short term in vitro maintained cytolytic T cell lines were shown to have mixed phenotypes by FACS analysis; approximately 50% to 75% of the cells in these populations were CD4-, CD8+, whereas 20% to 40% were CD4-, CD8-. Experiments in which effector cells were positively selected by adherence to anti-CD8 mAb-treated plates confirmed that CD8+ T cell could exhibit Ag-specific cytolytic activity against T. cruzi-infected target cells. Efforts are under way to clone these CTL to test their in vivo function and to determine their Ag specificity.