When neonatal pigs continuously fed formula are supplemented with leucine pulses, muscle protein synthesis and body weight gain are enhanced. To identify the responsible mechanisms, we combined plasma metabolomic analysis with transcriptome expression of the transcriptome and protein catabolic pathways in skeletal muscle. Piglets (n = 23, 7-day-old) were fed continuously a milk replacement formula via orogastric tube for 21 days with an additional parenteral infusion (800 μmol kg⁻¹ h⁻¹) of either leucine (LEU) or alanine (CON) for 1 h every 4 h. Plasma metabolites were measured by liquid chromatography-mass spectrometry. Gene and protein expression analyses of longissimus dorsi muscle were performed by RNA-seq and Western blot, respectively. Compared with CON, LEU pigs had increased plasma levels of leucine-derived metabolites, including 4-methyl-2-oxopentanoate, beta-hydroxyisovalerate, β-hydroxyisovalerylcarnitine, and 3-methylglutaconate (P ≤ 0.05). Leucine pulses downregulated transcripts enriched in the Kyoto Encyclopedia of Genes and Genomes terms “spliceosome,” “GAP junction,” “endocytosis,” “ECM-receptor interaction,” and “DNA replication”. Significant correlations were identified between metabolites derived from leucine catabolism and muscle genes involved in protein degradation, transcription and translation, and muscle maintenance and development (P ≤ 0.05). Further, leucine pulses decreased protein expression of autophagic markers and serine/threonine kinase 4, involved in muscle atrophy (P ≤ 0.01). In conclusion, results from our studies support the notion that leucine pulses during continuous enteral feeding enhance muscle mass gain in neonatal pigs by increasing protein synthetic activity and downregulating protein catabolic pathways through concerted responses in the transcriptome and metabolome.

当连续饲喂配方奶的新生猪补充亮氨酸豆类时，肌肉蛋白质合成和体重增加都会增强。为了确定相关机制，我们将血浆代谢组学分析与骨骼肌中转录组和蛋白质分解代谢途径的转录组表达相结合。仔猪（n  = 23，7 日龄）通过口胃管连续饲喂代乳配方奶 21 天，并额外进行肠外输注（800 μmol kg ^-1  h ^-1) 亮氨酸 (LEU) 或丙氨酸 (CON) 每 4 小时 1 小时。通过液相色谱-质谱法测量血浆代谢物。分别通过RNA-seq和Western印迹对背最长肌进行基因和蛋白质表达分析。与 CON 相比，LEU 猪血浆亮氨酸衍生代谢物水平升高，包括 4-methyl-2-oxopentanoate、beta-hydroxyisovalerate、β-hydroxyisovalerylcarnitine 和 3-methylglutaconate（P ≤ 0.05)。亮氨酸脉冲下调了京都基因和基因组百科全书术语“剪接体”、“间隙连接”、“内吞作用”、“ECM 受体相互作用”和“DNA 复制”中丰富的转录本。亮氨酸分解代谢产生的代谢物与参与蛋白质降解、转录和翻译以及肌肉维持和发育的肌肉基因之间存在显着相关性（P  ≤ 0.05）。此外，亮氨酸脉冲降低了与肌肉萎缩有关的自噬标志物和丝氨酸/苏氨酸激酶 4 的蛋白质表达（P ≤ 0.01)。总之，我们的研究结果支持这样的观点，即连续肠内喂养期间的亮氨酸脉冲通过增加蛋白质合成活性和通过转录组和代谢组中的协同反应下调蛋白质分解代谢途径来增强新生猪的肌肉质量增加。