L450W and Q455K Col8a2 Knock-In Mouse Models of Fuchs Endothelial Corneal Dystrophy Show Distinct Phenotypes and Evidence for Altered Autophagy

摘要

Purpose.: We compared the cellular phenotypes and studied the role of autophagy in the pathogenesis of Fuchs endothelial corneal dystrophy (FECD) using two ?±2 collagen VIII (Col8a2) knock-in mouse models and human FECD tissues. Methods.: In vivo corneal endothelial cell (CEC) counts and morphology were analyzed by clinical confocal microscopy. Ultrastructural analysis of CECs was performed by transmission electron microscopy. Real-time PCR and Western blotting were performed using total RNA, and protein extracted from mouse CECs and human CECs obtained from FECD and autopsy patients. Results.: Both Col8a2 mouse models exhibited hallmarks of FECD; however, the Col8a2 L450W/L450W mice exhibited a milder phenotype compared to the Col8a2 Q455K/Q455K mice. Both models exhibited upregulation of the unfolded protein response (UPR) as evidenced by dilated rough endoplasmic reticulum (RER), and upregulation of UPR-associated genes and proteins. Real-time PCR of Col8a2L450W/L450W and Col8a2Q455K/Q455K CECs at 40 weeks revealed a 2.1-fold (P 0.05) and a 5.2-fold (P 0.01) upregulation of the autophagy marker Dram1, respectively. Real-time PCR of human FECD endothelium revealed a 10.4-fold upregulation of DRAM1 (P 0.0001) compared to autopsy controls. Conclusions.: The Col8a2 L450W/L450W and Col8a2 Q455K/Q455K mouse models of FECD showed distinct endothelial cell phenotypes. Dram1 was associated with activation of the UPR and increased autophagy. Overexpression of this gene in mouse and human FECD endothelial cells suggested a role for altered autophagy in this disease.