BACKGROUND AND AIM OF THE STUDY: Aortic valve homografts were treated with 50% ethanol and glycerol followed by freeze-drying (D-Hydro). Comparative results of fresh versus D-Hydro-treated aortic roots implanted for up to nine months in the descending aorta of sheep with induced aortic regurgitation (AR) are reported. METHODS: Six fresh and six D-Hydro valves were implanted in 12 sheep for three, six and nine months, and echocardiography and pressures were taken at surgery and sacrifice. Tissue sections were stained with hematoxylin and eosin, von Kossa, Masson's trichrome, Movat's pentachrome, von Willebrand factor, CD3 (a T-cell marker) and smooth muscle alpha-actin. RESULTS: No grafts had increased gradients after implantation, or at sacrifice. At explantation, fresh homografts showed early pannus formation followed by thrombus, annular dilatation and wall calcification. Leaflets were thickened and progressively retracted. All had severe AR. The appearance of D-Hydro-treated homografts was normal, except for mild leaflet retraction in three, resulting in AR (in two animals the induced AR had healed). Histologically, a T-cell-mediated reaction was evident in the fresh homografts, and collagen distortion was noted. Calcification was present in all fresh specimens and was severe at nine months. D-Hydro roots showed only minor calcification in the six-month samples. Normal collagen, and a complete layer of von Willebrand factor-stained cells were present at three months. At nine months, cell rehabitation extended for two-thirds of the leaflets (alpha-actin +). The inflammatory reaction was very mild, with CD3+-stained cells absent in most samples. CONCLUSION: Aortic valve homografts treated with the D-Hydro freeze-drying method performed better than fresh homografts due to the absence of thrombus and annulus dilatation, limited calcification, and rehabitation of the aortic wall and parts of the leaflet by myofibroblasts, as well as the presence of a complete endothelial layer on the aortic wall and leaflet.