Extrarenal citrulline disposal in mice with impaired renal function

The endogenous synthesis of arginine, a semiessential amino acid, relies on the production of citrulline by the gut and its conversion into arginine by the kidney in what has been called the “intestinal-renal axis” for arginine synthesis. Although the kidney is the main site for citrulline disposal, it only accounts for ∼60–70% of the citrulline produced. Because the only known fate for citrulline is arginine synthesis and the enzymes that catalyze this reaction are widespread among body tissues, we hypothesized that citrulline can be utilized directly by tissues to meet, at least partially, their arginine needs. To test this hypothesis, we used stable and radioactive tracers in conscious, partially nephrectomized (½ and ⅚) and anesthetized acutely kidney-ligated mouse models. Nephrectomy increased plasma citrulline concentration but did not affect citrulline synthesis rates, thus reducing its clearance. Nephrectomy (⅚) reduced the amount of citrulline accounted for as plasma arginine from 88 to 42%. Acute kidney ligation increased the half-life and mean retention time of citrulline. Whereas the rate of citrulline conversion into plasma arginine was reduced, it was not eliminated. In addition, we observed direct utilization of citrulline for arginine synthesis and further incorporation into tissue protein in kidney-ligated mice. These observations indicate that a fraction of the citrulline produced is utilized directly by multiple tissues to meet their arginine needs and that extrarenal sites contribute to plasma arginine. Furthermore, when the interorgan synthesis of arginine is impaired, these extrarenal sites are able to increase their rate of citrulline utilization.

De novo synthesis is the main source of ornithine for citrulline production in neonatal pigs

Citrulline is an amino acid synthesized in the gut and utilized for the synthesis of the conditionally essential amino acid arginine. Recently, the origin of the ornithine utilized for citrulline synthesis has become a matter of discussion. Multiple physiological factors may have contributed to the differences found among different researchers; one of these is the developmental stage of the subjects studied. To test the hypothesis that during the neonatal period de novo synthesis is the main source of ornithine for citrulline synthesis, neonatal piglets were infused intravenously or intragastrically with [U-13C6]arginine, [U-13C5]glutamine, or [U-13C5]proline during the fasted and fed periods. [ ureido-15N]citrulline and [2H2]ornithine were infused intravenously for the entire infusion protocol. During fasting, plasma proline (13%) and ornithine (19%) were the main precursors for citrulline synthesis, whereas plasma arginine (62%) was the main precursor for plasma ornithine. During feeding, enteral (27%) and plasma (12%) proline were the main precursors for the ornithine utilized in the synthesis of citrulline, together with plasma ornithine (27%). Enteral proline and glutamine were utilized directly by the gut to produce ornithine utilized for citrulline synthesis. Arginine was not utilized by the gut, which is consistent with the lack of arginase activity in the neonate. Arginine, however, was the main source (47%) of plasma ornithine and in this way contributed to citrulline synthesis. In conclusion, during the neonatal period, the de novo pathway is the predominant source for the ornithine utilized in the synthesis of citrulline, and proline is the preferred precursor.

Enteral arginase II provides ornithine for citrulline synthesis

The synthesis of citrulline from arginine in the small intestine depends on the provision of ornithine. To test the hypothesis that arginase II plays a central role in the supply of ornithine for citrulline synthesis, the contribution of dietary arginine, glutamine, and proline was determined by utilizing multitracer stable isotope protocols in arginase II knockout (AII−/−) and wild-type (WT) mice. The lack of arginase II resulted in a lower citrulline rate of appearance (121 vs. 137 μmol·kg−1·h−1) due to a reduced availability of ornithine; ornithine supplementation was able to restore the rate of citrulline production in AII−/− to levels comparable with WT mice. There were significant differences in the utilization of dietary citrulline precursors. The contribution of dietary arginine to the synthesis of citrulline was reduced from 45 to 10 μmol·kg−1·h−1 due to the lack of arginase II. No enteral utilization of arginine was observed in AII−/− mice (WT = 25 μmol·kg−1·h−1), and the contribution of dietary arginine through plasma ornithine was reduced in the transgenic mice (20 vs. 13 μmol·kg−1·h−1). Dietary glutamine and proline utilization were greater in AII−/− than in WT mice (20 vs. 13 and 1.4 vs. 3.7 μmol·kg−1·h−1, respectively). Most of the contribution of glutamine and proline was enteral rather than through plasma ornithine. The arginase isoform present in the small intestinal mucosa has the role of providing ornithine for citrulline synthesis. The lack of arginase II results in a greater contribution of plasma ornithine and dietary glutamine and proline to the synthesis of citrulline.