431
For allograft rejection to occur, alloreactive T cells must become activated and migrate to the graft site. Interaction between integrin adhesion molecules and their ligands are believed to be important in both processes. The purpose of this study was to develop a model in which T cell activation and homing could be monitored independently in vivo We then tested the mode, using a peptide inhibitor of 4 integrin/fibronectin (FN) interactions thought to specifically inhibit T cell homing without affecting activation. Methods. The 2C T cell receptor transgenic provides a purified alloreactive CD8+ cytotoxic T cell (CTL) population (B6 background) with specificity for the Class 1 MHC (Ld) of the BALB/c mouse. To study allorejection in vivo, we used an adoptive transfer model of allograft rejection. Bb-scid recipients were made diabetic with streptozotocin and transplanted with BALB/c islets under the renal capsule. Islets in non-reconstituted recipients survived indefinitely. The adoptive transfer of 2 x 106 2C splenic CTL led to rejection within 14 days. CTL homing was defined as the presence of 2C+ T cells in the islet allograft both histologically and by flow cytometry of the graft 10 days after transplantation. CTL activation was defined as the phenotypic expression of activation markers (IL2-receptor and LFA-I) on ZC+ cells in reconstituted animals in viva 10 days after transplantation. Since naive 2C CTL express 4 integrin (as measured by flow cytometry), we chose to study 4/FN interactions in homing and activation using a specific pentapeptide inhibitor of 4/FN adhesion termed connecting segment 1 peptide (CSI-P) via continuous infusion Alzet pumps (15mg/kg/dx 14 days). Results. Prior to injection, the 2C T cells expressed very low, but detectable levels of IL2R and LFA-I In B6-scid mice who did not receive a BALB/c graft, adoptively- transferred 2C cells did not show phenotypic signs of activation (negative control). In B6-scid recipients of BALB/c islet allograft (positive control), the 2C cells in the spleen showed signs of activation (high expression of IL2R and LFA-1). In addition, the islet allograft was infiltrated with large numbers of 2C+ T cells. CS1-P completely blocked 2C CTL homing to the islet allograft at 10 days after transplantation. In contrast, CSI- P did not block 2C CTL activation either in vivo (phenotypic evidence of 2C activation was found in CS, -P treated B6-scid recipients of BALB/c islet grafts) or in vivo (2C CTL proliferation in response to BALB/c stimulator cells was not blocked by CSI -P). Importantly, CS1-P treatment significantly delayed the 2C-mediated rejection of BALB/c islet grafts (147+/-3.3 controls vs 29.5+/14.4, p<0.01). Conclusion: We conclude that this 2C adoptive transfer model of allograft rejection is useful in assessing both CTL activation and homing in viva These data suggest that4/FN interactions are critically important for CTL homing to the graft, but do not significantly affect activation. Thus, CS1 -P may represent a unique class of immunosuppressive agents that prolongs allograft survival by inhibiting CTL homing.