Abstract
Purpose :
Maintaining a tight blood-retinal barrier (BRB) is important for retinal visual function, which is composed of an inner BRB (iBRB) formed by tight junctions between endothelial cells (ECs) and an outer BRB (oBRB) tight junctions between retinal pigment epithelial cells (RPE). Loss of iBRB leads to diabetic retinopathy while disruption of the oBRB causes age-related macular degeneration. However, little is known about the similarity or difference in bioenergetic signature of these retinal barrier cells. This study therefore aims to measure mitochondrial and cytosolic bioenergetic pathways in retinal barrier cells.
Methods :
Human retinal endothelial cell (HREC) and human retinal pigment epithelium (APRE-19) were used to monitor the two main bioenergetic pathways; mitochondrial oxidative phosphorylation (OXPHOS) and glycolysis by measuring Oxygen Consumption Rate (OCR) and extracellular acidification rate (ECAR), respectively, using a XFe96 flux Analyzer. Furthermore, Mito Stress Test assays measured OCR at four levels: with no addition, and after adding oligomycin (Olig), FCCP, and rotenone to compare the six parameters of mitochondrial function between HRECs and ARPE-19, including basal OCR, ATP-linked OCR, proton leak OCR, maximal OCR, reserve capacity, and non-mitochondrial OCR. On the other hand, glycolysis stress test assays were used to measure ECAR at four levels: with or without glucose, Olig, and 2-deoxyglucose to compare the four glycolytic parameters between HREC and ARPE-19, including glycolytic flux, glycolytic capacity, glycolytic reserve capacity, and nonglycolytic ECAR. Statistical analysis was performed using Student t test and p<0.05 was considered significant.
Results :
Our results show that there is no significant difference between ECAR glycolytic flux between ARPE-19 and HRECs. However, significant differences in mitochondrial functions between these two types of cells exist. This is evident in that ARPE-19 has higher levels of basal OCR, ATP-linked OCR, proton leak OCR, maximal OCR, and reserve capacity than HREC. Non-mitochondrial OCR is similar between these two retinal cell types.
Conclusions :
Retinal EC has different bioenergetics than RPE in that retinal EC relies more on glycolysis while RPE depends on OXPHOS. The results suggest that reprogramming metabolic signatures would be a new approach to treat different retinal diseases.
This is a 2020 ARVO Annual Meeting abstract.