Long‐term Leucine and BCAA Inclusion in a 30% Protein and Energy Restricted Diet Increases mTORC1 Signaling in Skeletal Muscle of Neonatal Pigs

Abstract

Suboptimal nutrient intake during the shift from parenteral to oral feeding represents a limiting factor for growth of low‐birth weight infants. Leucine has been shown to have anabolic properties in skeletal muscle and can promote lean gain when supplemented to neonatal pigs fed a milk replacement formula. The objective of this study was to test if enteral leucine supplementation alone, or in combination with isoleucine and valine, can increase muscle protein synthesis in neonate pigs who consume 30% less crude protein (CP) and metabolizable energy than their requirements, and whether this effect is mediated through changes in muscle protein synthesis and degradation pathways, and amino acid (AA) transporter expression. Piglets (4‐d‐old) were fed by gastric tube 1 of 4 diets providing (kg BW/d) 15 g CP and 215 kcal (CON), 10.5 g CP and 156 kcal (R), 10.5 g CP + 1.19 g leucine and 156 kcal (RL), or 10.5 g CP + 1.67 g branched‐chain amino acids and 156 kcal (RBCAA) at 4‐h intervals for 9 d (n = 24). Body weights (BW) were recorded on d 0, 3, 6, and 9. On d 9, plasma AA and insulin concentrations and muscle protein synthesis rate, abundance of Na+‐coupled neutral AA transporter (SNAT2) and L‐type AA transporter (LAT1), and phosphorylation and/or abundance of insulin receptor substrate 1 (IRS‐1, Ser1101), protein kinase B (PKB), S6 kinase (S6K1), 4E‐binding protein (4EBP1), eukaryotic initiation factor 4 complex (eIF4E·eIF4G), forkhead box O (FOXO1, Ser256), atrophy F‐Box/Atrogin‐1 and muscle RING‐finger protein (MuRF1) were determined post‐feeding. Plasma leucine levels were greater in RL and RBCAA compared to CON (P ≤ 0.01). BW gain and muscle protein synthesis rate, phosphorylation of S6K1 and 4EBP1, and formation of eIF4E·eIF4G were lower in R, RL, and RBCAA than CON (P < 0.01). RBCAA and RL upregulated (P ≤ 0.01) S6K1 and 4EBP1 phosphorylation and eIF4E·eIF4G formation compared to R. Phosphorylation of FOXO1 was lower in R compared to CON (P ≤ 0.05). There were no differences in phosphorylation and/or abundance of IRS‐1, PKB, SNAT2, LAT1, atrogin‐1 and MURF1 among diets. In conclusion, when protein and energy intakes are restricted, leucine or BCAA supplementation for 9 d increases the activation of the mammalian target of rapamycin pathway, but does not enhance skeletal muscle protein synthesis or promote body weight gain in neonatal pigs. Furthermore, protein degradation signaling, AA transporter abundance, and insulin signaling are unaffected by leucine or BCAA supplementation of a protein and energy restricted diet suggesting that when both dietary protein and/or energy supplies are deficient, the anabolic effect of leucine or BCAA supplementation on muscle protein accretion is limited.Support or Funding InformationNIH AR444474, NIH HD072891, NIH AR46308, USDA NIFA 2013‐67015‐20438, and USDA/ARS 6250‐51000‐055