Multiple approaches in studying the biochemical effects of amino acid supplementation in preruminant calves

Resumen

As the muscle system is one of the most fast-growing compartments in calves before weaning, it requires a large amount of protein deposition. Amino acids, as the basic unit of proteins, play crucial roles in the growth of animals. Supplementation of several amino acids in milk replacers has been used in calf industry for decades and several approaches have been established to assess the functionality, outcome and efficiency of this supplementation. In this work, a “multi-omics” approach has been used, aiming to use large scale of data collected from metabolomic and proteomic analyses (known as “multi-omics” or “foodomics”) to elucidate the impact of several amino acid supplementations in the milk replacer for calves. The work of this project is divided into three studies and “multi-omics” were applied in Study 1 and Study 2: In Study 1, proteomics and metabolomics (“multi-omics”) and serum clinical chemistry were used to study the skeletal muscle of calves under supplementation with several amino acids in milk replacers. Animals were divided into four groups: Ctrl, control diet without supplementation; GP, control supplemented with Glycine (G) and Proline (P); FY, control supplemented with phenylalanine (F) and tyrosine (Y); and MKT, control supplemented with methionine (M), lysine (K) and threonine (T). The integrated results indicated that MKT supplementation yielded a mild improvement of growth, increased serum creatinine and insulin, muscle Phospho-S6 / S6 ratio as well as some proteins related to muscle function, despite some of the elevations are non-significant. The metabolome analysis from skeletal muscle biopsies revealed several differences between the GP - FY groups and the Ctrl - MKT groups, suggesting a metabolic adaptation especially in GP and FY groups. In Study 2, the same approaches to the Study 1 were used, but both plasma and skeletal muscle biopsies were included in the “multi-omics” analysis. Animals were divided into three groups: Ctrl, control diet without supplementation; LT, control supplemented with leucine (L) and threonine (T); and Leu, control supplemented with leucine. In both supplemented treatments, the integrated results indicated that leucine and its metabolites were available in the organism. In the LT group compared to the Ctrl group, integrated metabolomics and proteomics showed that branched-chain amino acids (BCAA) degradation and mitochondrial oxidative metabolism (citrate cycle, respiratory chain) were the main activated pathways in muscle. Energy metabolic substrates would be BCAA derivatives, butyrate from ruminal production and/or lipid mobilization. The deleterious effects of activated oxidative phosphorylation were balanced by the upregulation of antioxidant proteins. An increase in protein synthesis was indicated by the increase in Phospho-S6 / S6 ratio in skeletal muscle. In the Leu group, animals suffered digestive problems at the beginning of the study and the experimental design was not correctly randomized when biopsies were obtained. The serum/plasma chemical, proteomic and metabolomic analyses showed a profile compatible with a pathological process and a biopsy-caused bias. In Study 3, reference intervals for several biochemical parameters for unweaned calves and recently weaned piglets at different ages were calculated using large number of animals sampled at different ages from populations under different season trials. The main variable was age whereas no major trial- or sex-biased differences were noticed. This information will help veterinarians and animal science researchers to better practice their professions by providing them with more adequate diagnostic references.