Defined Regional Analysis of Mitochondrial Activity in the Endothelium of Donor Corneal Tissue

Abstract

Purpose : To obtained a detailed metabolic activity profile and comparison among defined regions of the corneal endothelium.

Methods : Corneoscleral tissues from donors 50-75 years old were marked at the temporal sclera during recovery. Endothelium-Descemet membrane complex (EDM) punches obtained from central, temporal, superior, nasal, and inferior regions of 14 corneas (11 donors) were assayed for mitochondrial activity (oxygen consumption rate [OCR; pmole/min]) using extracellular flux analysis of oxygen and compared. Mitochondrial density from 20 additional corneas (20 donors) was compared among regions using a mitochondria-selective dye and fluorescence imaging.

Results : Comparing raw OCR/cell values of peripheral to central endothelium, only temporal samples at baseline displayed a difference (P=0.02). OCR measurements for remaining peripheral samples compared to central endothelium did not differ (P>0.16). Differences in baseline OCR/cell between central and temporal samples were associated with greater ATP production in temporal samples (P=0.04). Comparing raw OCR/cell values among peripheral samples, only temporal and superior regions displayed differences at baseline (P=0.02) and post-oligomycin (P=0.04). Differences in baseline and post-oligomycin OCR/cell between temporal and superior samples were associated with greater proton leak in temporal samples (P=0.06). Agreement plots comparing OCR/cell between central and peripheral regions suggest nasal samples most closely approximate central endothelial samples. In the mitochondrial density analysis, no differences were detected among any of the regions.

Conclusions : Extracellular flux analysis of oxygen consumption using tissue from the either the superior, nasal, or inferior endothelial regions provides a valid approximation of central metabolic activity. Results provide a foundation for using small peripheral corneal endothelium samples to investigate the relationship between metabolic activity and physiological factors influencing tissue function.

This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.