Effects Of Leucine Supplementation On Amino Acids Metabolism And Protein Synthesis In Skeletal Muscle Of Holstein Calves

The high rate of protein synthesis in skeletal muscle of Holstein calves can benefit their first lactation even lifetime milk yield.Protein turnover has been ongoing in skeletal muscle,which decomposes old proteins and re-synthesizes new proteins,during which essential amino acids(EAA)from protein breakdown are partially oxidatively decomposed and lost.Reducing EAA loss and making full use of EAA for skeletal muscle protein synthesis are of great significance for calves' growth.Studies have found that leucine,one of the functional amino acids,is a proteogenic AA that also functions as a signaling molecule to stimulate translational initiation through m TOR pathway.Therefore,we hypothesized that the leucine supplementation can promote translation initiation by activating the m TOR pathway,competitively attracting more EAA into the translation process,thereby promoting protein synthesis.Ten male neonatal Holstein calves(38 ± 3 kg)were randomly assigned to either the control(CON,n=5)or supplmentation with 1.435 g leucine /L milk(LEU,n=5).Within one hour after birth,all calves were fed 4.0 L colostrum and then housed in individual calf hutch.Days 2 to 6 were adaption period,when the amounts of alanine and leucine added into the milk were increased by 20% daily till 100% on day 6.Starter feed was provided from week 3.Calves were fed with pasteurized milk and starter feed twice daily at 08:00 and 17:00.The amount of the diet was adjusted weekly to ensure all feed was consumed without any refusal.Fresh drinking water was available all the time.The aims of this study were to determine the effects of leucine supplementation on fasted AA metabolism including connection with EAA utilization and protein synthesis,and further explore whether protein synthesis in skeletal muscle of Holstein calves can be improved.The results and conclusions are as follows:1.Supplementation of leucine increased the concentration of methionine,threonine,histidine and EAAs(P < 0.05)and decreased the concentration of arginine in serum(P < 0.05).Treatment × time interactions were observed on serine,histidine and glycine(P < 0.05).All amino acids in serum had a time effect(P < 0.01),and the concentrations of BCAA,EAA,non-essential amino acids(NEAA),and total amino acid(TAA)in the first week were higher than those in other periods(P < 0.05).Compared with the control group,the concentrations of free isoleucine and hydrolyzed phenylalanine in the skeletal muscle in the leucine group were lower(P < 0.05).What's more,leucine supplementation decreased level of creatinine and GOT activity(P < 0.05),and showed a tendency to reduce the GPT activity(P = 0.062)in skeletal muscle.In conclusion,the supplementation of leucine inhibited the transamination in the skeletal muscle and maintained relatively high EAA concentration of Holstein calves.2.Compared to the control group,leucine supplementation significantly reduced the concentration of pyruvate acid in the liver(P < 0.05),increased the concentrations of glutamate,acetyl-Co A and glucose in the liver(P < 0.05),and also improved the glucose concentration in the blood(P < 0.05).As a result,the supplementation of leucine increased the available energy of Holstein calves.3.Compared to the control group,leucine supplementation significantly enhanced the phosphorylation of mammalian target of rapamycin(m TOR),eukaryotic initiation factor 4E binding protein 1(4EBP1)and ribosomal protein S6 kinase 1(p70S6K),and no significant difference was determined in the phosphorylation of e IF2?(P > 0.05).In conclusion,the supplementation of leucine stimulated protein synthesis and skeletal muscle development of Holstein calves by activating the m TOR and its downstream signaling molecules 4EBP1 and S6K1.The results of this studies indicated that leucine supplementation increased the available EAA and energy of Holstein calves,and promotes translation initiation by activating m TOR and its downstream signaling molecules S6K1 and 4EBP1,thereby competitively utilizing EAA to promote protein synthesis in skeletal muscle and its development of Holstein calves.