Elucidating the molecular basis of PPCD: Effects of decreased ZEB1 expression on corneal endothelial cell function.

Purpose: To investigate the functional role that the () gene, which underlies the genetic basis of posterior polymorphous corneal dystrophy 3 (PPCD3), plays in corneal endothelial cell proliferation, apoptosis, migration, and barrier function.

Methods: A human corneal endothelial cell line (HCEnC-21T) was transfected with siRNA targeting mRNA. Cell proliferation, apoptosis, migration, and barrier assays were performed: Cell proliferation was assessed with cell counting using a hemocytometer; cell apoptosis, induced by either ultraviolet C (UVC) radiation or doxorubicin treatment, was quantified by measuring cleaved caspase 3 (cCASP3) protein levels; and cell migration and barrier function were monitored with electric cell-substrate impedance sensing (ECIS).

Results: knockdown in HCEnC-21T cells transfected with siRNA targeting did not result in a significant difference in cell proliferation when compared with control. Although knockdown of in HCEnC-21T cells sensitized the cells to UV-induced apoptosis, knockdown did not alter the cells' susceptibility to doxorubicin-induced apoptosis, as measured with cCASP3 protein levels, compared with controls. Similarly, no difference was observed in cell migration following knockdown. However, cell barrier function increased significantly following knockdown.

Conclusions: The corneal endothelium in PPCD3 is characterized by morphologic, anatomic, and molecular features that are more consistent with an epithelial-like rather than an endothelial-like phenotype. Although these characteristics have been well documented, we demonstrate for the first time that susceptibility to UV-induced apoptosis and cell barrier function are significantly altered in the setting of reduced . The significance of an altered cellular response to apoptotic stimuli and increased cell barrier function in the pathobiology of PPCD remains to be fully elucidated.