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Lithium treatment increases endothelial cell survival and autophagy in a mouse model of Fuchs endothelial corneal dystrophy.

ABSTRACT: Lithium previously has been shown to reduce both endoplasmic reticulum (ER) and oxidative stress in other in vitro and in vivo model systems. We investigated lithium's effects on cultured corneal endothelial cells (CECs) exposed to these types of stress and in a mouse model of Fuchs endothelial corneal dystrophy (FECD).Viability of cultured bovine CECs was determined by CellTiter-Glo. 2-month-old Col8a2(Q455K/Q455K) mutant (Q455K) and C57/Bl6 wild type animals were divided into two groups of 15 mice. Group I received 0.2% lithium carbonate-containing chow and Group II received control chow for 7 months. Confocal microscopy, transmission electron microscopy, real-time PCR (RT-PCR) and western blot were performed.Pretreatment with lithium increased viability of cultured CECs after H2O2 and thapsigargin exposure compared with untreated controls (p<0.05). In vivo analysis of mouse corneal endothelium showed the following: endothelial cell density of lithium treated Q455K was higher than for untreated Q455K (p<0.01). transmission electron microscopy of lithium treated Q455K showed normal endothelium with enlarged autophagosomes, but untreated Q455K showed dilated ER and guttae. Compared with untreated Q455K endothelium, lithium treated Q455K showed significant upregulation of P62, Tmem74, Tm9sf1 and Tmem166 by RT-PCR and of Atg5-12 conjugate by western blotting indicating that lithium treatment increased autophagy. Although RT-PCR unexpectedly showed increased levels of lithium response genes, caspase 12, Gsk3?, Arr?2 and Impa1, western blotting showed the expected downregulation of Arr?2 and Impa1 proteins in response to lithium treatment.Lithium increases cultured CEC survival against ER and oxidative stress. Increased autophagy in lithium treated endothelium in a mouse model of FECD suggests autophagy may contribute to increased endothelial cell survival.

SUBMITTER: Kim EC

PROVIDER: S-EPMC3793892 | biostudies-literature | 2013 Aug

REPOSITORIES: biostudies-literature

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Lithium treatment increases endothelial cell survival and autophagy in a mouse model of Fuchs endothelial corneal dystrophy.

Kim Eun Chul EC Meng Huan H Jun Albert S AS

The British journal of ophthalmology 20130612 8

<h4>Background</h4>Lithium previously has been shown to reduce both endoplasmic reticulum (ER) and oxidative stress in other in vitro and in vivo model systems. We investigated lithium's effects on cultured corneal endothelial cells (CECs) exposed to these types of stress and in a mouse model of Fuchs endothelial corneal dystrophy (FECD).<h4>Methods</h4>Viability of cultured bovine CECs was determined by CellTiter-Glo. 2-month-old Col8a2(Q455K/Q455K) mutant (Q455K) and C57/Bl6 wild type animals ...[more]

PMID: 23759441

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Project description:Purpose To develop a model to predict corneal improvement after Descemet membrane endothelial keratoplasty (DMEK) for Fuchs endothelial corneal dystrophy (FECD) from Scheimpflug tomography. Design Cross-sectional study. Participants Forty-eight eyes (derivation group) and 45 eyes (validation group) with a range of severity of FECD undergoing DMEK. Methods Scheimpflug images were obtained before and after DMEK. Before DMEK, pachymetry map and posterior elevation map patterns were quantified by a special image analysis program measuring tomographic features of edema (loss of regular isopachs, displacement of the thinnest point of the cornea, posterior surface depression). Image-derived novel parameters were combined with instrument-derived parameters, and the relative influences of parameters associated with the change in central corneal thickness (CCT) after DMEK in the derivation group were determined by using a gradient boosting machine learning model. The parameters with highest relative influence were then fit in a linear regression model. The derived model was applied to the validation group. Correlations and agreement were assessed between predicted and observed changes in CCT. Main Outcome Measures Predictive power (R2) and mean difference between predicted and observed change in CCT. Results The gradient boosting machine model identified 4 novel parameters of isopach circularity and eccentricity and 1 instrument-derived parameter (posterior surface radius); preoperative CCT was a poor predictor. In the derivation group, the model strongly predicted the change in CCT after DMEK (R2 = 0.80; 95% confidence interval [CI], 0.71–0.89) and the mean difference between predicted and observed change was, by definition, 0 μm. When the same 5 parameters were fit to the validation group, the model performed very highly (R2 = 0.89; 95% CI, 0.84–0.94). When the coefficient estimates from the derivation model were used to predict the change in CCT in the validation group, the predictive power was also high (R2 = 0.78; 95% CI, 0.68–0.88), and the mean difference was 4 μm (predicted minus observed). Conclusions Scheimpflug tomography maps of corneas with FECD can predict the improvement in CCT after DMEK, independent of preoperative corneal thickness measurement. The model could be applied in clinical practice or for clinical research of FECD.

Project description:Fuchs’ endothelial corneal dystrophy (FECD) is a progressive vision impairing disease caused by thickening of Descemet’s membrane and gradual degeneration and loss of corneal endothelial cells. The aim of this study was to identify differentially expressed genes between FECD-affected and unaffected corneal endothelium to gain insight into the pathophysiological mechanisms underlying this disease. Microarray gene expression analysis was performed on total RNA from FECD-affected and unaffected corneal endothelium-Descemet’s membrane (CE-DM) specimens using the Illumina HumanHT-12 v3.0 expression array. RNA from pools of FECD-affected (n=3 per pool) and individual unaffected (n=3) specimens was used for comparison. Altered expression of a sub-set of differentially expressed genes was validated by quantitative reverse transcription-polymerase chain reaction (qRT-PCR) in independent specimens. Bioinformatics analysis was performed using InnateDB to reveal functional relationships among the differentially expressed genes and molecular pathways involved in the disease. A total of 16,513 genes were found expressed in the corneal endothelium of which 142 genes were differentially expressed between FECD-affected and unaffected endothelium (log2 fold-change ≥1.5, corrected p-value ≤0.05). Most of the genes were up-regulated (126) and a small proportion down-regulated (16) in affected corneal endothelium. Of the twelve genes prioritised for validation, differential expression of 10 genes, including those ranked 57th and 81st by significance validated by qRT-PCR (8 up-regulated and 2 downregulated, corrected p ≤0.05), one gene showed a trend for up-regulation in affected endothelium, consistent with the microarray analysis and another was up-regulated in an independent study indicating robustness of the differential expression dataset. Bioinformatic analysis revealed significant over-representation of differentially expressed genes in extracellular matrix reorganisation, cellular remodelling, immune response, and inflammation. Network analysis showed functional inter-relatedness of the majority of the dysregulated genes and revealed known direct functional relationships between 20 of the genes; many of these genes have roles in macrophage differentiation, phagocytosis and inflammation. This is the second report of microarray gene expression analysis in FECD. This study revealed a set of highly dysregulated genes in the corneal endothelium in FECD. More than a third of the dysregulated genes in the disease have been discovered for the first time and thus are novel. The dysregulated genes strongly suggest the presence of phagocytic cells, most likely immune cells, and inflammation in corneal endothelium in the disease. This study provides a molecular framework for delineating the mechanisms underlying these cellular processes in FECD.

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Lithium treatment increases endothelial cell survival and autophagy in a mouse model of Fuchs endothelial corneal dystrophy.

Publications

Lithium treatment increases endothelial cell survival and autophagy in a mouse model of Fuchs endothelial corneal dystrophy.

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