Abstract
Purpose :
To study the mechanism underlying the development of age-related cataract and disrupted fiber cell morphogenesis and calcium homeostasis in connexin 46 knockout (Cx46KO) lenses at 129S4(129) and C57BL/6J(B6) strain backgrounds. To determine how cell-cell junctional complex, cytoskeleton, and their associated scaffold proteins such as periaxin (Prx) are involved in the regulation of fiber cell shape and surface interlocking structures and in maintaining the lens homeostasis.
Methods :
Mouse lens phenotypes were characterized by slit lamp examination and imaging analysis. Fiber cell morphogenesis was characterized by immunostaining with specific markers and high-resolution confocal imaging. Lens protein changes and protein-protein interactions were evaluated by western blotting, co-labeling with specific markers or mass-spec analysis. Calcium deposits were examined using alizarin red dye staining.
Results :
Confocal imaging results showed disruptions of cell shape, surface interlocking structures and cytoskeletons in lens deep cortical regions where had no obvious opacity, but severely deformed fibers with aberrant membrane/cytoskeletal aggregates in the core of 129-Cx46KO lens with severe nuclear cataract. The 129-Prx proteins were detected in fiber cell protrusions and colocalized with F-actin in vivo and in vitro. A loss of periaxin prevented severe nuclear cataracts in 129-Cx46KO PrxKO (DKO) lenses. Many 129-Prx-associated junctional proteins were identified. Alizarin red dye staining revealed calcium deposits in the lens suture regions of 129-Cx46KO, B6-Cx46KO and DKO lenses. Cleaved alpha-crystallins were detected in 129-Cx46KO, B6-Cx46KO and DKO lens homogenates.
Conclusions :
Periaxin is important in the regulation of membrane/cytoskeletal complexes to control fiber cell morphogenesis. 129-Prx promotes the formation of light-scattering aggregates that consist of membrane/cytoskeletal protein complexes in 129-Cx46KO nuclear cataracts. Increased calcium deposits alone are associated with mild cataracts in lens suture regions of both DKO and B6 Cx46KO lenses. This work reveals novel molecular insights about the roles of cell junctions/scaffold protein linkers/cytoskeletal superstructural complexes in fiber cell morphogenesis in the lens formation and age-related cataractogenesis.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.