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Extensive tracheal defects may pose a life-threatening dilemma. Although tracheal transplantation may represent a reconstructive solution, very little is known regarding the immunobiology and behavior of tracheal allografts. The objective of this study was to assess the pattern and kinetics of re-epithelialization of orthotopic tracheal allografts in immunosuppressed recipients. Thirty-eight age-matched mice were randomly assigned to five experimental groups. BALB/c donor tracheal segments were orthotopically transplanted into either syngeneic BALB/c or MHC mismatched allogeneic C57BL/6 recipients with and without immunosuppression. On post-transplant days 7, 14, 28, 48, and 62, animals from each group were evaluated by serial histology, electron microscopy, and serial immunohistochemical analysis for mucosal phenotype, re-epithelialization pattern, and lymphocyte subpopulations. Nonimmunosuppressed recipients underwent recipient-derived basal cell re-epithelialization by Day 48, with differentiation into a sparse population of ciliated columnar epithelium by Day 62, whereas immunosuppressed recipients underwent basal cell re-epithelialization 28 d after transplantation and differentiation into a dense population of ciliated columnar epithelium by Day 48. The re-epithelialization process occurred in a definable pattern that was significantly enhanced with the addition of immunosuppression. Orthotopic tracheal transplants undergo progressive re-epithelialization with recipient-derived basal cells that differentiate into ciliated columnar epithelium in a definable pattern that is enhanced with the addition of immunosuppression.