Excess glutamine rewires endothelial cell metabolism.

Abstract

Introduction: Glutamine, the most abundant amino acid in the body, is a key metabolic substrate for endothelial cells. Glutamine supplementation protects against cardiovascular disease in animal models and in humans; however, glutamine in vitro has inconsistent effects on endothelial function. Furthermore, little is known about how altered metabolite concentrations, for example excess glucose in hyperglycemia or excess glutamine in cell culture media, affect endothelial cell metabolism.

Objectives: The objective of this study was to determine how physiological and supplemented glutamine affect endothelial metabolism in normal and high glucose conditions.

Methods: Primary human coronary artery endothelial cells were cultured in varied glutamine concentrations and in normal and high glucose. Glutamine uptake and glutamate secretion were measured using a YSI bioanalyzer; oxidative respiration was assessed using a Seahorse Metabolic Analyzer; and glutamine carbon incorporation into the TCA cycle, amino acids, antioxidants, and other pathways was evaluated via liquid chromatography-mass spectrometry.

Results: As extracellular glutamine increased, endothelial cells took up more glutamine, but glutamate secretion saturated above 2 mM glutamine. Excess glutamine was primarily stored intracellularly, although increasing extracellular glutamine concentration did increase oxidative respiration and TCA cycle isotope enrichment. We also observed increased glutamine incorporation into glutathione, UDP-GlcNAc, and amino acids. When total metabolite abundance was examined, intracellular succinate, unsaturated fatty acids, and one-carbon metabolism-related metabolites decreased with increasing glutamine.

Conclusion: These findings demonstrate that excess extracellular glutamine reprograms endothelial metabolism, suggesting that glutamine supplementation should be used with caution in cardiovascular therapies and endothelial cell culture.