Islet rejection in perforin-deficient mice: the role of perforin and Fas.

BACKGROUND

Perforin and Fas are the two main pathways by which cytotoxic T lymphocytes (CTLs) mediate target cell lysis in vitro. The perforin pathway is predominantly used by CD8+ cells, which comprise the majority of CTLs. The Fas pathway has been demonstrated to be the principal cytolytic mechanism in CD4+ CTLs. CTLs have been shown to play an important role in allograft rejection. In this study, we examined the relevance of perforin and Fas to allograft rejection by transplanting pancreatic islets from fully allogeneic C3H/HeJ (C3H) or Fas-deficient C3H/lpr donors into perforin-deficient (P0) mice or controls with intact perforin genes (P2).

METHODS

P0 or P2 mice that were rendered diabetic with streptozotocin at 300 mg/kg i.p. received approximately 350 islets obtained from C3H or C3H/lpr donors by in situ collagenase digestion and Ficoll density centrifugation of the pancreas. Four groups of animals were studied: C3H to P2 (group 1), C3H to P0 (group 2), lpr to P0 (group 3), and syngeneic P2 to P2 (group 4). Graft survival monitored by blood sugar levels was compared among the groups. At the time of rejection (blood sugar >300 mg/100 ml), grafts were harvested and analyzed by histopathology, immunocytochemistry, and reverse transcriptase-polymerase chain reaction. Primary splenic T cells of the recipients, harvested at the time of rejection, were tested for cytotoxicity against 51Cr-labeled donor cells.

RESULTS

The mean graft survival for groups 1, 2, and 3 was 10.2+/-1.4, 12.2+/-6.0, and 13.2+/-0.8 days, respectively. Syngeneic grafts survived indefinitely. Rejecting grafts from all groups (1, 2, and 3) showed an intense infiltration by both CD4+ and CD8+ cells and complete islet destruction. Reverse transcriptase-polymerase chain reaction revealed granzyme B in rejecting grafts from all three groups.

CONCLUSIONS

Perforin and Fas pathways alone or in combination are not required for islet rejection, suggesting that these pathways may not play a crucial role in allograft rejection.