Maternal supplementation with citrulline or arginine during gestation impacts fetal amino acid availability in a model of intrauterine growth restriction (IUGR).

BACKGROUND: Supplementing maternal diet with citrulline or arginine during gestation was shown to enhance fetal growth in a model of IUGR induced by maternal dietary protein restriction in the rat. OBJECTIVE: The aims of this study were to determine in the same model whether maternal supplementation with citrulline or arginine would increase 1) citrulline and arginine concentration in fetal circulation; 2) the expression of placental amino acid transporters, and 3) the fetal availability of essential amino acids. METHODS: Pregnant rats (n = 8 per group) were fed either an isocaloric control (20% protein, NP) or a low protein (LP, 4% protein) diet, either alone or supplemented with 2 g/kg/d of l-citrulline (LP + CIT) or isonitrogenous Arginine (LP + ARG) in drinking water throughout gestation. Fetuses were extracted by C-section on the 21st day of gestation. The gene expression of system A (Slc38a1, Slc38a2, and Slc38a4) and L (Slc7a2, Slc7a5, Slc7a8) amino acid transporters was measured in placenta and amino acid concentrations determined in maternal and fetal plasma. RESULTS: Maternal LP diet decreased fetal (4.01 ± 0.03 vs. 5.45 ± 0.07 g, p < 0.0001) and placental weight (0.617 ± 0.01 vs. 0.392 ± 0.04 g, p < 0.001), by 26 and 36% respectively, compared with NP diet. Supplementation with either CIT or ARG increased fetal birth weight by ≈ 5 or 11%, respectively (4.21 ± 0.05 and 4.48 ± 0.05 g vs. 4.01 ± 0.03 g, p < 0.05). CIT supplementation produced a 5- and 2-fold increase in fetal plasma citrulline and arginine, respectively, whereas ARG supplementation only increased fetal arginine concentration. LP diet led to lower placental SNAT 4 mRNA, and higher LAT2 and SNAT1 expression, compared with NP. SNAT4, 4hFC, LAT2 mRNA were up-regulated in LP + CIT and LP + ARG group compared with the un-supplemented LP group. Higher level of LAT1 mRNA was also observed in the LP + CIT group than in the LP group (p < 0.01). SNAT2 expression was unchanged in response to CIT or ARG supplementation. Fetal amino acid concentrations were decreased by LP diet, and were not restored by CIT or ARG supplementation. CONCLUSIONS: The current findings confirm supplementation with citrulline or arginine enhances fetal growth in a rat model of IUGR. They further suggest that: 1) citrulline and arginine administered orally to the pregnant mother may reach fetal circulation; 2) citrulline effectively raises fetal arginine availability; and 3) although it failed to increase the concentrations of essential amino acids in fetal plasma, citrulline or arginine supplementation upregulates the gene expression of several placental amino acid transporters.

Maternal citrulline supplementation enhances placental function and fetal growth in a rat model of IUGR: involvement of insulin-like growth factor 2 and angiogenic factors.

OBJECTIVE: To determine the effects of maternal citrulline supplementation on fetal growth and placental efficiency in a rat model of intrauterine growth restriction (IUGR) induced by maternal protein restriction. METHODS: Pregnant Sprague-Dawley rats were randomly assigned to three groups: NP (receiving a control 20% protein diet), LP (a 4% protein diet), or LP-CIT (an LP diet along with L-citrulline, 2 g/kg/d in drinking water). On the 15th and 21st day of gestation (GD15 and GD21, respectively), dams underwent a C-section, by which fetuses and placentas were extracted. The expression of genes involved in placental growth and angiogenesis was studied by quantitative RT-PCR. RESULTS: Maternal citrulline supplementation increased fetal weight at GD21, and fetal weight/placental weight ratio, an index of placental efficiency, from mid gestation (p < 0.001). The expression of Igf2-P0, a placenta-specific variant of insulin-like growth factor 2 (Igf2) gene, and Vegf and Flt-1, involved in angiogenic pathways, was enhanced in the LP-CIT group (versus NP, p < 0.001, p < 0.01, and p < 0.05 for Igf2-P0, Vegf, and Flt-1, respectively). CONCLUSIONS: In a model of IUGR induced by protein deprivation, citrulline enhances fetal growth, placental efficiency, and the expression of genes involved in angiogenesis. The relevance of such effect in human pregnancies complicated by IUGR warrants further study.

L-Citrulline Supplementation Enhances Fetal Growth and Protein Synthesis in Rats with Intrauterine Growth Restriction.

BACKGROUND: Intrauterine growth restriction (IUGR) results from either maternal undernutrition or impaired placental blood flow, exposing offspring to increased perinatal mortality and a higher risk of metabolic syndrome and cardiovascular disease during adulthood. l-Citrulline is a precursor of l-arginine and nitric oxide (NO), which regulates placental blood flow. Moreover, l-citrulline stimulates protein synthesis in other models of undernutrition. OBJECTIVE: The aim of the study was to determine whether l-citrulline supplementation would enhance fetal growth in a model of IUGR induced by maternal dietary protein restriction. METHODS: Pregnant rats were fed either a control (20% protein) or a low-protein (LP; 4% protein) diet. LP dams were randomly allocated to drink tap water either as such or supplemented with l-citrulline (2 g · kg(-1) · d(-1)), an isonitrogenous amount of l-arginine, or nonessential l-amino acids (NEAAs). On day 21 of gestation, dams received a 2-h infusion of l-[1-(13)C]-valine until fetuses were extracted by cesarean delivery. Isotope enrichments were measured in free amino acids and fetal muscle, liver, and placenta protein by GC-mass spectrometry. RESULTS: Fetal weight was ∼29% lower in the LP group (3.82 ± 0.06 g) than in the control group (5.41 ± 0.10 g) (P < 0.001). Regardless of supplementation, fetal weight remained below that of control fetuses. Yet, compared with the LP group, l-citrulline and l-arginine equally increased fetal weight to 4.15 ± 0.08 g (P < 0.05) and 4.13 ± 0.1 g (P < 0.05 compared with LP), respectively, whereas NEAA did not (4.05 ± 0.05 g; P = 0.07). Fetal muscle protein fractional synthesis rate was 35% lower in the LP fetuses (41% ± 11%/d) than in the control (61% ± 13%/d) fetuses (P < 0.001) and was normalized by l-citrulline (56% ± 4%/d; P < 0.05 compared with LP, NS compared with control) and not by other supplements. Urinary nitrite and nitrate excretion was lower in the LP group (6.4 ± 0.8 µmol/d) than in the control group (17.9 ± 1.1 µmol/d; P < 0.001) and increased in response to l-citrulline or l-arginine (12.1 ± 2.2 and 10.6 ± 0.9 µmol/d; P < 0.05), whereas they did not in the LP + NEAA group. CONCLUSION: l-Citrulline increases fetal growth in a model of IUGR, and the effect may be mediated by enhanced fetal muscle protein synthesis and/or increased NO production.