Abstract
Purpose :
Autosomal recessive congenital hereditary endothelial dystrophy type 2 (CHED2) is associated with either homozygous or compound heterozygous mutations in SLC4A11. Corneal edema is observed at early infancy. The Slc4a11 KO mouse is a good model for studying the pathophysiology of CHED2. With the goal of characterizing this model we study five parameters at early (12 weeks) and late (40 weeks) age: 1) Corneal thickness, 2) Corneal endothelial density, 3) Corneal endothelial permeability, 4) Tight junction and actin cytoskeleton structure, 5) Oxidative stress.
Methods :
Corneal thickness was evaluated by Spectral-Domain Optical Coherence Tomography in vivo. Corneal endothelial density was quantified by ZO-1 immunostaining of corneal buttons followed by counting. Corneal endothelial permeability was evaluated by mounting whole corneas in small cups and adding 0.1% Na-Fluorescein on the endothelial side. After incubation and wash, fluorescence was quantified. Corneal buttons were stained by MitoSox and endothelial immunofluorescence was performed for ZO-1 and F-actin. Hmox1 was quantified by QPCR and immunostaining.
Results :
In Slc4a11 KO mice corneal thickness was increased 90% at 12 weeks (WT: 95.0±9.2, KO: 181.0±24.8 μm, p=1x10-9, n=9) and 202% at 40 weeks (WT: 104.5±8.4, KO: 315.56±20.49 μm, p=9x10-5, n=9). Corneal endothelial density was not different at 12 weeks (WT: 2566±108, KO: 2457±125 Cells/mm2, p=0.2, n=3) and reduced only 19% at 40 weeks (WT: 1872±42, KO: 1518±89 Cells/mm2, p=9x10-4, n=3), however hexagonal morphology was lost. Corneal endothelial permeability to fluorescein increased 23% in KO at 12 weeks of age (WT: 5413±243, KO: 6663±120 rfu, p=0.01, n=2). Tight junctions were partially disrupted as ribbon-like structure of ZO-1 was replaced by a line pattern and F-actin cytoskeletal rings associated with cell junctions were disordered at 40 weeks of age. These changes are consistent with compromised barrier function in corneal endothelium. Markers of oxidative stress MitoSox and Hmox1 were increased at early and late age.
Conclusions :
Slc4a11 KO mice show a similar phenotype as CHED2; that is severe corneal edema at early age with no changes in corneal endothelial cell density. Corneal endothelial barrier function is compromised at early and late age. Oxidative stress may be related to the pathological process as markers are increased at early age.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.