Connexin 50 and AQP0 are Essential in Maintaining Organization and Integrity of Lens Fibers

Abstract

PurposeConnexins and aquaporins play essential roles in maintaining lens homeostasis and transparency and there is a close physical and functional relationship between these two proteins. Aquaporin 0 (AQP0), in addition to its role in water transport in the lens, acts as a cell–cell adhesion molecule. Recently, we showed a new role of connexin (Cx) 50 in mediating cell–cell adhesion. However, the cooperative roles of these two proteins in the lens in vivo have not been reported.MethodsWe generated an AQP0/Cx50 double knockout (dKO) mouse model. Light, fluorescence, transmission thin section, and freeze-fracture electron microscopy, as well as wheat germ agglutinin and phalloidin labeling were used to evaluate lens structure. Mechanical properties of lenses were determined by mechanical compression testing.ResultsDKO mice exhibited small eyes and lenses with severe cataracts, along with lens posterior defects, including posterior capsule rupture. The dKO mouse lenses had severe structural disruption associated with increased spaces between lens fiber cells when compared with wild-type lenses or lenses deficient in either Cx50 or AQP0. DKO mice also exhibited greater reduction in lens size compared with Cx50 KO mice. Gap-junction plaque size was greatly decreased in cortical fiber cells in dKO mice. Moreover, lens stiffness and elasticity were completely diminished, exhibiting a gelatinous texture in adult dKO mice.ConclusionsThis novel mouse model reveals that Cx50 and AQP0 play an important role in mediating cell–cell adhesion function in the lens fiber cells and their deficiency impairs lens fiber organization, integrity, mechanical properties, and lens development.

Macrophage recruitment in immune-privileged lens during capsule repair, necrotic fiber removal, and fibrosis

“…This is more evident in the posterior region of the lens, including posterior subcapsular and polar cataracts, and posterior capsular rupture with severe extrusion of lens fiber cells. We also reported disorganized fiber cell structures with abnormally enlarged extracellular spaces ( Gu et al., 2019 ). By using this dKO mouse model, we report a direct mechanism for macrophages in the immune-privileged lens organ during damage repair and fibrosis.…”

“…Our previous studies have shown a significant reduction in the eyeball and lens size in Cx50KO and Cx50/AQP0 dKO mice compared with wild-type (WT) and AQP0KO mice ( Gu et al., 2019 ). Here, we found that the increase in lens mass and diameter of the dKO was delayed during postnatal (P) development, and the growth rate was significantly less than those of WT and single KO, especially compared with WT and AQP0 KO groups ( Figures S1 A and S1 B).…”

“…The posterior capsule ruptured phenotypes are also reported in mutant Lop10/Lop10 α3 −/− mice ( Chang et al., 2002 ; Xia et al., 2006 ). In our dKO mouse model, the posterior capsule rupture is primarily attributed to a reduced cell-adhesion function with severe exacerbation of lens fiber cell organization and excessive enlargement of extracellular spaces as a result of Cx50/AQP0 deficiency in lens fibers ( Hu et al., 2017 ; Gu et al., 2019 ). These severe disruptions of lens fiber structure are likely to generate extra physical stress on the thin, vulnerable posterior capsule, given the posterior capsule is thinner than that in other parts of the lens, and its thickness is decreased with aging ( Barraquer et al., 2006 ; Krag and Andreassen, 2003 ).…”

“…Similar cataract phenotypes are also observed in Cx46, Cx50, AQP0 knockout (KO) or mutated gene knock-in mouse models ( White et al., 1998 ; Al-Ghoul et al., 2003 ; Lo et al., 2014 ; Gong et al., 1997 ). By generating an AQP0/Cx50 double-KO (dKO) mouse model, we show that Cx50 and AQP0 play important roles in mediating cell-cell adhesion in lens fibers, and their deficiency impairs fiber organization, integrity, mechanical properties, and lens development ( Gu et al., 2019 ). Moreover, lens fiber cells in dKO mice “leak” out of the lens through the ruptured posterior capsule.…”

Regulation of Connexin Gap Junctions and Hemichannels by Calcium and Calcium Binding Protein Calmodulin

“…The intracellular loop and C-terminal domains are highly diverse among connexin subtypes [6]. Three connexins present in the lens are Cx43, Cx46, and Cx50, of which Cx46 and Cx50 are predominantly expressed in the lens fiber cells [7]. Epithelial cells predominantly express Cx43 and Cx50, while Cx50 and Cx46 are present in bulk of lens fiber cells.…”