Energy substrate requirements of rat retinal pigmented epithelial cells in culture: relative importance of glucose, amino acids, and monocarboxylates.
OBJECTIVE
To determine the metabolic conditions that provide maintenance of viability for cultured rat RPE cells and to determine whether monocarboxylates such as lactate or pyruvate, which are known to exist at high concentrations in the subretinal space, can provide an alternative energy source to maintain cells when other nutritive supplies are limited.


METHODS
Cultured rat RPE cells (passage 2-4), in the absence of serum, were subjected to different metabolic challenges relating to glucose, amino acid, or oxygen deprivation. Lactate or pyruvate was added to some cells in each instance to determine whether cells could be maintained by using these substances as fuel sources for metabolic reactions. Cell viability was assessed after treatments, and in some cases proliferation rates and appearance of apoptosis-like DNA cleavage were also investigated by terminal deoxynucleotidyl-transferase UTP-linked nick-end labeling (TUNEL). Western blot analysis was used to determine the expression of transporters for glucose and monocarboxylates in these cells.


RESULTS
RPE cell viability was partially reduced in the absence of glucose or with glycolytic inhibition. Lactate or pyruvate did not prevent these reductions. Inhibition of transaminase reactions with aminooxyacetic acid (AOAA) in the absence of glucose caused a complete loss of viability that was reversed by pyruvate or lactate. MCT inhibition was detrimental to RPE cell viability only at high concentrations (500 microM) in the presence of glucose but blocked the protective effect of pyruvate-lactate in the presence of AOAA at 1 microM.


CONCLUSIONS
Rat RPE cells require glucose as their primary metabolic substrate in culture, but can metabolize glutamine in its absence. When glucose and glutamine are limiting, RPE cells can metabolize monocarboxylates such as lactate or pyruvate. These data provide evidence that such cells are able to withstand various types of insult brought about by nutrient deprivation, by altering their pathways of energy production.
Wood
John P M
JP
Nuffield Laboratory of Ophthalmology, University of Oxford, Oxford, United Kingdom.
Chidlow
Glyn
G
Graham
Mark
M
Osborne
Neville N
NN
D000596:Amino Acids / Q000378:metabolism*; D000818:Animals; D015153:Blotting, Western; D018929:Cell Culture Techniques; D002455:Cell Division; D002470:Cell Survival; D004247:DNA / Q000378:metabolism; D004734:Energy Metabolism; D005947:Glucose / Q000378:metabolism*; D015151:Immunoblotting; D007124:Immunoenzyme Techniques; D020287:In Situ Nick-End Labeling; D019344:Lactic Acid / Q000378:metabolism*; D027501:Monocarboxylic Acid Transporters / Q000378:metabolism; D009004:Monosaccharide Transport Proteins / Q000378:metabolism; D010857:Pigment Epithelium of Eye / Q000166:cytology / Q000378:metabolism*; D019289:Pyruvic Acid / Q000378:metabolism*; D051381:Rats
D000596:Amino Acids; D027501:Monocarboxylic Acid Transporters; D009004:Monosaccharide Transport Proteins; D019344:Lactic Acid; D019289:Pyruvic Acid; D004247:DNA; D005947:Glucose