Mitochondrial Cell Death Pathway in Ocular Anterior Segment Development

Abstract

Purpose: : Anterior segment dysgenesis (ASD) is a complex congenital ocular disorder which affects the normal development of most of the anterior segment tissues in the eye. Peters’ anomaly is a common form of ASD, which manifests during early stage of eye development and is characterized by the presence of a lens stalk. Children with Peters’ anomaly have a cloudy central cornea and often develop cataract and glaucoma. It has been implicated that programmed cell death might play an important role in lens-corneal separation. In this study, we tested the hypothesis in mouse models that activation of the classical mitochondrial cell death pathway is essential for the lens vesicle to detach from the surface ectoderm during early anterior segment development.

Methods: : Two apoptosis-defective mouse models were used in the study. One is the cytochrome c (Cyt-C) knock-in mice in which the normal Cyt-C is replaced by a mutant protein (CytC-KA) that is incapable of activating the downstream target protein Apaf-1. The other mouse model is the Apaf-1 knockout mice. Developmental defects in the anterior segments were examined by histology. Cell death in mouse embryonic eyes were analyzed by TUNEL assay and immunofluorescence for cleaved (active) caspase-3.

Results: : At embryonic day 16.5 (E16.5), both CytC-KA knock-in (CytC^KA/KA) and Apaf-1^-/- embryos developed typical features of Peter’s anomaly including presence of a lens stalk, adhesion of lens to the cornea, partial or complete absence of the corneal endothelium and anterior chamber. At E11.5, TUNEL assay showed that the number of apoptotic cells in the anterior zone of the lens vesicle was reduced significantly or eliminated in the CytC^KA/KA mutant and Apaf1^-/- mouse embryos respectively, in comparison to the age-matched CytC^KA/+ and Apaf-1^+/- embryos. In consistence with the TUNEL result, immunoreactivity of active caspase-3, a key effector caspase activated in the mitochondrial cell death pathway, was also reduced or absent in the lens vesicle of the mutant mice.

Conclusions: : We demonstrated that blocking the mitochondrial cell death pathway in mouse results in anterior segment dysgenesis which resembles Peters’ anomaly in humans. Our results suggest that activation of mitochondrial cell death pathway is required for the lens vesicle to detach from the surface ectoderm. Our study implies that pathogenesis of Peters’ anomaly could result from dysfunction of the mitochondrial apoptosis machinery during early eye development.

Keywords: anterior segment • apoptosis/cell death • mitochondria