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Liver transcriptomic and proteomic profiles of preweaning lambs are modified by milk replacer restriction

ABSTRACT: Purpose: The present study was designed to identify both differentially expressed (DE) genes and differences in proteins accumulated in the liver tissues of suckling female lambs, thus trying to identify modified metabolic pathways as a consequence of milk restriction during the suckling period. Methods: Forty Assaf lambs (average BW 4.7 kg) were penned individually, twenty of them were fed milk replacer (200 g dry matter/L) ad libitum (ADL; 192 mL/kg LBW) whereas the other group (restricted, RES) only received 120 mL/kg LBW. When they were 35 days old, four animals per group were slaughtered (8 lambs in total) and a piece of liver was excised for transcriptomic analysis. The liver transcriptome analysis was carried out using RNA sequencing methodology (RNA-seq). Results: 386 DE genes were identified by RNA-seq, 198 of them being annotated genes in the KEGG pathway. Positive values of log2-fold change (log2FC) indicated that 210 genes were up-regulated in the liver of RES relative to the ADL group, whereas negative log2FC values denoted the down-regulation of 176 genes (P < 0.05). Conclusions: According to the data obtained, a restricted milk intake during the suckling period of replacement lambs affects hepatic transcriptome and proteome associated with an altered metabolism of lipids and proteins, thus reducing feed efficiency of replacement period.

ORGANISM(S): Ovis aries

PROVIDER: GSE110192 | GEO | 2018/12/16

REPOSITORIES: GEO

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Project description:Feed restriction during the early life of ewe lambs (F0) triggers the transfer of epigenetic marks to the next generation, thus impairing the reproductive performance of F1. However, the effects of this factor on milk production and composition, including its abundance in regulatory miRNA (many of which are contained in exosomes, small vesicles of endocytic origin that play a role in the modulation of immune response of the offspring), has not been tested so far. Therefore, in this study, the replacement ewe lambs (F0) obtained in a previous project (a group of ewes fed milk replacer ad libitum –ADL– vs. a group of ewes restricted –RES– to 62.5% the intake level of milk replacer during the suckling period) were raised under similar post-weaning conditions and mated to obtain the progeny (F1). The F1 dairy ewes were also mated, and six of these F1 dairy ewes were selected and divided into two groups: the F1-ADL group (n=3) born from F0-ADL and the F1-RES group (n=3), born from F0-RES animals. Milk production was controlled during the peak lactation period, and milk samples were obtained for each gland separately to measure chemical composition, somatic cell counts, and bacteriology. Exosomes were also isolated for each gland separately to define the miRNA content. The miRNAome was determined following a Next Generation Sequencing approach. The first preliminary results indicate significant differences in the abundance of five miRNAs in the milk obtained during the peak of lactation. Thus, oar-miR-150, oar-miR-221, oar-miR-23a, oar-miR-27a, oar-miR-376c were all down modulated in F1-RES when compared to F0-ADL. Most of these miRNAs have been found to play a role in biological functions such as development, apoptosis, muscle differentiation, reproduction, or milk production. Moreover, somatic cell count (SCC) was reduced significantly in milk samples during the peak of lactation of F1-RES dairy sheep. No significant differences were found in milk production, the chemical composition of milk (fat, protein, lactase), or bacteriology (colony forming units, CFU). This study's results provide some evidence of the effects of nutritional programming events on the milk's bioactive components.

Project description:The common practise of artificially rearing some lambs from prolific meat breeds of sheep constitutes a welfare issue due to increased mortality rates and negative health issues. In this multidisciplinary study, we investigated the possible short and mid-term advantages of artificially feeding fresh ewe’s milk instead of commercial milk replacer on lambs’ growth, health and welfare. Romane lambs were either separated from their mothers on D3 and fed with Lacaune ewes’ milk (LAC, n=13) or milk replacer (REP, N=15), or they were reared by their mothers (MOT, n=15). On D45, they were weaned, gathered in single sex groups until the end of the study on D150. Lamb performance and biomarkers of overall health were assessed by measuring: growth, dirtiness of the perianal area, enteric pathogens in the faeces, total antioxidant status and redox status assessed by plasma reduced (GSH)/oxidized (GSSG) glutathione ratio, and immune response after vaccination against chlamydiosis. As an exploratory approach, blood cell transcriptomic profiles were also investigated. Last, Qualitative Behaviour Assessment was performed as an integrated welfare criteria. LAC and REP never differed in their average daily gain but grew less than MOT lambs in the early suckling period and just after weaning. No effect was detected afterwards. On D30, LAC and REP lambs had lower total antioxidant and higher redox status than MOT lambs but did not differ among themselves. LAC and MOT had a cleaner perianal area than REP lambs on D21, while faecal pathogen infection did not vary between the treatment groups. After vaccination, LAC also had a stronger immune response on D90 compared to REP lambs. Transcriptome analysis performed on D150 showed differential gene expression, mainly in relation to inflammatory, immune and cell cycle response, between male lambs of the LAC group and those of the MOT and REP groups. Based on Qualitative Behaviour Assessment, LAC lambs never differed from MOT lambs in their general activity and varied from REP only on D21; REP lambs were always more agitated than MOT lambs. In conclusion, artificial milk feeding impaired early growth rate, health, and emotional state mainly during the milk feeding period and at weaning. Feeding artificially reared lambs with fresh ewe's milk partly mitigated some of the negative effects induced by milk replacer but without achieving the full benefit of being reared by the mother.

Project description:A summary of the work associated to these microarrays is the following: Diet plays a role in the development of the immune system, and polyunsaturated fatty acids can modulate the expression of a variety of genes. Human milk contains conjugated linoleic acid (CLA), a fatty acid that seems to contribute to immune development. Indeed, recent studies carried out in our group in suckling animals have shown that the immune function is enhanced after feeding 80:20 isomer mix of c9,t11 and t10,c12 CLA, respectively. However, little work has been done on the effects of CLA on gene expression, and even less regarding immune system development in early life. With all these concepts in mind, we determined the expression profile of Mesentheric Lymph Nodes (MLN) from animals supplemented with CLA during gestation and suckling through dam’s milk (Group A) or by oral gavage (Group B), supplemented just during suckling (Group C) and control animals (Group D) with the aid of the specific GeneChip ® Rat Genome 230 2.0 (Affymettrix). Bioinformatic analyses were performed using the GeneSpring GX software package v10.0.2 and lead to the identification of 123 genes differentially expressed in all three dietary approaches. Generation of a Biological Association Network (BAN) evidenced several genes, such as connective tissue growth factor (Ctgf), tissue inhibitor of metalloproteinase 1 (Timp1), galanin (Gal), synaptotagmin 1 (Syt1), growth factor receptor bound protein 2 (Grb2) actin gamma 2 (Actg2) and smooth muscle alpha actin (Acta2), as highly interconnected nodes of the resulting network. Gene underexpression was confirmed by Real-Time RT-PCR. We conclude that Ctgf, Timp1, Gal and Syt1, among others, are genes modulated by CLA supplementation that may have a role on lymphoproliferation and mucosal immune responses in early life. The aim of our study was to evaluate, by using whole genome microarrays, the effects of dietary supplementation with an 80:20 isomer mix of c9,t11 and t10,c12 CLA, respectively, on Mesentheric Lymph Nodes gene expression, during gestation and/or suckling. Three experimental approaches were conducted to assess the effects of CLA supplementation on I) pups from dams fed with 1% CLA diet during the two weeks of gestation and throughout the suckling period (Group A). These pups received CLA through the dam’s milk during suckling (Total Period of CLA Supplementation (TPS) 5 wk); II) pups from dams fed with 1% CLA diet during gestation and standard diet during suckling. These pups were CLA-supplemented daily during suckling by oral gavage (TPS 5 wk) (Group B); III) pups from dams fed standard diet during gestation and suckling and receiving CLA by daily oral gavage throughout the suckling period (TPS 3 wk) (Group C). Group D, pups from dams fed standard diet throughout the study, constituted the reference diet group (TPS 0 wk). Triplicate samples were hybridized for each experimental condition (12 samples in total). The samples provided were analyzed using the specific software GeneSpring GX.

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Liver transcriptomic and proteomic profiles of preweaning lambs are modified by milk replacer restriction

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