Microcirculatory Function during Endotoxemia-A Functional Citrulline-Arginine-NO Pathway and NOS3 Complex Is Essential to Maintain the Microcirculation.

Competition for the amino acid arginine by endothelial nitric-oxide synthase (NOS3) and (pro-)inflammatory NO-synthase (NOS2) during endotoxemia appears essential in the derangement of the microcirculatory flow. This study investigated the role of NOS2 and NOS3 combined with/without citrulline supplementation on the NO-production and microcirculation during endotoxemia. Wildtype (C57BL6/N background; control; n = 36), Nos2-deficient, (n = 40), Nos3-deficient (n = 39) and Nos2/Nos3-deficient mice (n = 42) received a continuous intravenous LPS infusion alone (200 µg total, 18 h) or combined with L-citrulline (37.5 mg, last 6 h). The intestinal microcirculatory flow was measured by side-stream dark field (SDF)-imaging. The jejunal intracellular NO production was quantified by in vivo NO-spin trapping combined with electron spin-resonance (ESR) spectrometry. Amino-acid concentrations were measured by high-performance liquid chromatography (HPLC). LPS infusion decreased plasma arginine concentration in control and Nos3-/- compared to Nos2-/- mice. Jejunal NO production and the microcirculation were significantly decreased in control and Nos2-/- mice after LPS infusion. No beneficial effects of L-citrulline supplementation on microcirculatory flow were found in Nos3-/- or Nos2-/-/Nos3-/- mice. This study confirms that L-citrulline supplementation enhances de novo arginine synthesis and NO production in mice during endotoxemia with a functional NOS3-enzyme (control and Nos2-/- mice), as this beneficial effect was absent in Nos3-/- or Nos2-/-/Nos3-/- mice.

Citrulline Supplementation Improves Organ Perfusion and Arginine Availability under Conditions with Enhanced Arginase Activity.

Enhanced arginase-induced arginine consumption is believed to play a key role in the pathogenesis of sickle cell disease-induced end organ failure. Enhancement of arginine availability with L-arginine supplementation exhibited less consistent results; however, L-citrulline, the precursor of L-arginine, may be a promising alternative. In this study, we determined the effects of L-citrulline compared to L-arginine supplementation on arginine-nitric oxide (NO) metabolism, arginine availability and microcirculation in a murine model with acutely-enhanced arginase activity. The effects were measured in six groups of mice (n = 8 each) injected intraperitoneally with sterile saline or arginase (1000 IE/mouse) with or without being separately injected with L-citrulline or L-arginine 1 h prior to assessment of the microcirculation with side stream dark-field (SDF)-imaging or in vivo NO-production with electron spin resonance (ESR) spectroscopy. Arginase injection caused a decrease in plasma and tissue arginine concentrations. L-arginine and L-citrulline supplementation both enhanced plasma and tissue arginine concentrations in arginase-injected mice. However, only the citrulline supplementation increased NO production and improved microcirculatory flow in arginase-injected mice. In conclusion, the present study provides for the first time in vivo experimental evidence that L-citrulline, and not L-arginine supplementation, improves the end organ microcirculation during conditions with acute arginase-induced arginine deficiency by increasing the NO concentration in tissues.

L-citrulline improves splanchnic perfusion and reduces gut injury during exercise.

PURPOSE: Splanchnic hypoperfusion is a physiological phenomenon during strenuous exercise. It has been associated with gastrointestinal symptoms and intestinal injury and may hamper athletic performance. We hypothesized that L-citrulline supplementation improves splanchnic perfusion and decreases intestinal injury by enhancing arginine availability. The aim of this study was to determine the effect of L-citrulline intake on splanchnic perfusion, intestinal injury, and barrier function during exercise. METHODS: In this randomized, double-blind crossover study, 10 men cycled for 60 min at 70% of their maximum workload after L-citrulline (10 g) or placebo (L-alanine) intake. Splanchnic perfusion was assessed using gastric air tonometry. Sublingual microcirculation was evaluated by sidestream dark field imaging. Plasma amino acid levels and intestinal fatty acid binding protein concentrations, reflecting enterocyte damage, were assessed every 10 min. Urinary excretion of sugar probes was measured to evaluate intestinal permeability changes. RESULTS: Oral L-citrulline supplementation enhanced plasma citrulline (1840.3 ± 142.3 µM) and arginine levels (238.5 ± 9.1 µM) compared with that in placebo (45.7 ± 4.8 µM and 101.5 ± 6.1 µM, respectively, P < 0.0001), resulting in increased arginine availability. Splanchnic hypoperfusion was prevented during exercise after L-citrulline ingestion (reflected by unaltered gapg-apCO2 levels), whereas gapg-apCO2 increased with placebo treatment (P < 0.01). Accordingly, L-citrulline intake resulted in an increased number of perfused small sublingual vessels compared with that in placebo (7.8 ± 6.0 vs -2.0 ± 2.4, P = 0.06). Furthermore, plasma intestinal fatty acid binding protein levels were attenuated during exercise after L-citrulline supplementation compared with that in placebo (AUC0-60 min, -185% ± 506% vs 1318% ± 553%, P < 0.01). No significant differences were observed for intestinal permeability. CONCLUSIONS: Pre-exercise L-citrulline intake preserves splanchnic perfusion and attenuates intestinal injury during exercise in athletes compared with placebo, probably by enhancing arginine availability. These results suggest that oral L-citrulline supplementation is a promising intervention to combat splanchnic hypoperfusion-induced intestinal compromise.