Isotope-guided metabolomics dissects kidney arginine metabolism

Abstract

Background: L-arginine is a key amino acid for detoxification, protein metabolism, and kidney and cardiovascular health. In the urea cycle, ammonia is incorporated into urea through the transformation of arginine to ornithine and urea, a reaction that occurs mostly in the liver but also in the kidneys. Hypothesis: An imbalance of kidney arginine metabolism can be associated with kidney injury. Aim: To detect the arginine uptake and its metabolic fate in kidney in vivo and ex vivo models by metabolic flux analysis. Method: Healthy C57BL/6 mice (n=22) were fed with normal and 13C6-15N4-arginine diet for 1, 2 and 8 weeks. Extracted metabolites from kidney cortex and medulla were analyzed with UHPLC/QQQ-based mass spectrometry targeting arginine-related metabolic pathways: the urea cycle, polyamines, the nitric oxide pathway, arginine/proline/glutamate interconversion, as well as modified forms of arginine. Additionally, micro-dissected nephron segments (glomeruli, proximal convoluted tubules (PCT), proximal straight tubules (PST), thin ascending limb of the loop of Henle (TAL), distal convoluted tubules (DCT), collecting ducts (CD)) from 4 healthy C57BL/6 mice, were incubated with 13C6-arginine and analyzed accordingly. Results: Isotope tracing allowed us to follow the fate of arginine in the kidney cortex and medulla of the 13C6-15N4-arginine-fed animals. The ex vivo experiments confirmed that all nephron segments take up and metabolize arginine, except for the TAL. More specifically, ornithine was found in all segments apart from the CD. Agmatine was formed in the PST and the CD, whereas arginine contributed to the proline and glutamine formation in the glomeruli, the PCT and the CD. Methylated forms of arginine were observed in the PST and the DCT. Conclusion: This study enabled the detection of arginine-related metabolites inside the kidney and showed that the metabolic fate of arginine differs along the nephron. The detailed isotope-based arginine metabolic flux and its role in kidney disease models will be determined next. This work is supported by the Young Investigator Award from the Novo Nordisk Foundation (grant number NNF19OC0056043), awarded to Markus Rinschen. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.