Prior to grafting, cell morphology was evaluated in vitro by phase-contrast microscopy (TS100 Eclipse; Nikon, Melville, NY, USA). Corneal endothelial cells seeded on glass coverslips were fixed (paraformaldehyde 4% for 30 minutes) for immunolabeling. Cells were first labeled with Alexa Fluor 488 phalloïdin (Invitrogen) for 30 minutes, then immunostained for 1 hour with either a mouse anti-sodium-potassium adenosine triphosphatase alpha 1 (Na+/K+-ATPase α1) (Millipore, Etobicoke, ON, Canada), a polyclonal anti-zonula occludens-1 (ZO-1) (Invitrogen), a mouse anti-alpha-smooth muscle actin (α-SMA) (Dako, Burlington, ON, Canada), a goat anti-fibronectin (Santa Cruz, Dallas, TX, USA), or a mouse anti-collagen type I (Sigma-Aldrich Corp.). The secondary antibodies Alexa Fluor 594 goat anti-mouse (Invitrogen), chicken anti-goat (Invitrogen), and chicken anti-rabbit (Invitrogen) were used as secondary antibodies for 45 minutes at room temperature. Antibodies were diluted in phosphate-buffered solution containing 1% bovine albumin serum (Sigma-Aldrich Corp.). Hoechst reagent 33258 or 4′,6-diamidino-2-phenylindole (DAPI) was added for cell nuclei counterstaining. Fluorescence was then observed (Axio Imager.Z2 microscope; AxioVision Rel. 4.8.2.; Carl Zeiss Canada, Toronto, ON, Canada), and the percentage of α-SMA–positive cells was determined (ImageJ; U.S. National Institutes of Health, Bethesda, MD, USA).