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Chun-hong Xia, Nikki Tjahjono, Rachel Li, Sarah Chu, Xiaohua Gong; Characterization of epithelial cell proliferation and differentiation in connexin mutant mice. Invest. Ophthalmol. Vis. Sci. 2019;60(9):4294.
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© ARVO (1962-2015); The Authors (2016-present)
Mouse Cx50-R205G is a point mutation located in the extracellular loop 2 (E2) of Gja8 or connexin 50 (Cx50), it is homologous to Cx50-R198W in humans, a mutation linked to cataract-microcornea syndrome. This project aims to investigate how Gja8 mutations including null mutation (Cx50-/-) and point mutation (Cx50-R205G) influence lens epithelial cell (LEC) proliferation and fiber cell differentiation to perturb lens growth and transparency. Cx50-/- mice develop small lenses with mild cataracts while Cx50-R205G mutant mice display much smaller lenses with severe nuclear cataracts, we will elucidate molecular and cellular evidence to address the underlying mechanism.
LEC proliferation in wild-type and Cx50 mutant mice will be characterized and compared by EdU labeling, followed by whole lens imaging; immunostaining and confocal imaging of primary cultured lens epithelial cells and lens sections are performed to study the distribution of gap junction proteins. Single cell gene expression profiles will be used to determine the clustering of lens epithelial cells.
The Cx50-R205G mutant line has a semi-dominant inheritance pattern, the heterozygous mice have reduced lens size and mild cataracts, while homozygous mice have much smaller lenses and dense nuclear cataracts, as well as vacuole-like structures in lens cortical fibers. Differing from Cx50-/- mice, the Cx50-R205G mutation disrupts lens size and transparency early in development, and causes decreased and aberrant distribution of proliferating LECs in the germinative zone (GZ) in addition to decreased central epithelium (CE) proliferation. We observe reduced connexin plaque formation in differentiating cells in vitro and in vivo, as well as disrupted fiber cell differentiation in Cx50-R205G mutant lenses. Furthermore, single cell gene expression data of wild-type and mutant lenses reveal a drastic multiple cell cluster change.
Molecular signaling mediated by Cx50 gap junction communication is critical for maintaining and regulating cell senescence, proliferation and differentiation of lens epithelial cells. Cx50 null or R205G point mutants distinctively alter connexin gap junction formation to perturb sub clusters of epithelial cells to affect anterior epithelial cell proliferation and equatorial cell differentiation processes in vivo.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.
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