Muc4/sialomucin complex (SMC), a large heterodimeric mucin composed of an extracellular mucin subunit ASGP-1 and a transmembrane subunit ASGP-2, is present at the rat ocular surface localized mainly to the most superficial layers of the epithelia. To investigate corneal homeostasis and the functions of Muc4/SMC at the ocular surface, we developed a corneal epithelial cell culture system from corneal explants, from which migrating cells formed an epithelial sheet resembling the native epithelium with regard to microanatomy, expression of characteristic markers, cell migration, and Muc4/SMC expression. Cells migrating from the explants expressed smooth muscle actin. Proliferation was detected only on the edge of epithelial sheet in the immature epithelium and throughout the sheet in confluent cultures. Microscopy revealed that the epithelial sheet was formed from four to six layers of cells expressing keratin 3 and Muc4/SMC in forms identical to those expressed at ocular surface in vivo. Electron microscopy showed cells in various morphological states in the process of releasing from the surface of the multilayer (desquamating). Surprisingly, few of these cells showed evidence of apoptosis, either by morphological or DNA fragmentation analyses. These results suggest a new model for desquamation from stratified epithelia, in which desquamation and apoptosis are independent and sequential processes. Desquamating cells also exhibit a high level of Muc4/SMC. Since Muc4/SMC has been shown to be a potent anti-adhesive and a repressor of apoptosis, we propose that it plays a role in the non-apoptotic desquamation process. © 2005 Wiley-Liss, Inc.