Dependency of the Trans Vivo Delayed Type Hypersensitivity Response on the Action of Regulatory T Cells: Implications for Monitoring Transplant Tolerance

Background. The trans vivo delayed-type hypersensitivity (DTH) assay has been used for monitoring the immune status of clinical transplant recipients. Here we tested the hypothesis that the assay can reveal control of allograft rejection by CD25 + CD4 + regulatory T cells (Treg). Methods. CBA.Ca (H2 k ) recipients ofheterotopic C57BL/10 (H2 b , B 10) heart transplants were untreated or pretreated with anti-CD4 antibody and donor-specific blood. This protocol has been shown previously to induce operational tolerance to alloantigens that is dependent on CD25 + CD4 + Treg. Four weeks after transplantation leukocytes were harvested and used for the trans vivo DTH assay. Cells were stimulated with irradiated B10 leukocytes or subcellular antigen and injected into ear pinnae of immune deficient CB17.SCID.beige hosts. Results. Stimulation of leukocytes from recipients rejecting B10 cardiac allografts with recall alloantigen caused a "strong" swelling response, whereas similar stimulation of leukocytes from operationally tolerant mice resulted in significantly less swelling (n=17; P=0.003). When CD25 + T cells were depleted from "tolerant" leukocytes, the swelling response triggered was similar to that obtained using cells from rejecting animals (P