OMICS APPROACHES FOR OMEGA-6/OMEGA-3 POLYUNSATURATED FATTY ACID RATIO IN PIGS

In general, meat quality is heavily influenced by fatty acid (FA) composition and intramuscular fat, which is mostly dependent on the dietary fat intake of pigs, but it is also affected by genetic variants. In this PhD thesis, we elucidated the effects of dietary lipids, in terms of omega-6 (n-6)/omega-3 (n-3) polyunsaturated fatty acid (PUFA) ratio, on porcine meat quality and the integration of OMICS technologies such as genomics, transcriptomics, and proteomics. With the aim of understanding the association between n-6/n-3 PUFA ratios, porcine transcriptome expression, and biological processes related to PUFA metabolism, we performed RNA-Seq and microRNA-Seq on longissimus dorsi muscle samples from 20 Iberian x Duroc pigs with extreme values for n-6/n-3 FA ratio and identified differentially expressed messenger RNAs (mRNAs) and microRNAs (miRNAs). Our findings highlighted mRNA genes, miRNAs and enriched pathways that were related to lipid metabolism, cell growth and inflammation, according to differences in muscle n-6/n-3 PUFA ratio. Relevant miRNA-to-mRNA regulatory networks were also uncovered (i.e., mir-15b to ARRDC3; mir-7142-3p to METTL21C), and linked to lipolysis, obesity, myogenesis, and protein catabolism. We also determined on how sow diets with high or low n-6/n-3 PUFA ratios of 13:1 (SOY) and 4:1 (LIN), influenced the reproductive performance of sows, growth performance of pre-weaned piglets, and fat deposition, including proteins and protein-coding genes using proteomics analysis. In sows, LIN decreased dead-born piglets and pre-weaning mortality. Although piglet weight and weight gain varied in between n-6/n-3 PUFA ratios, weaning weight and overall weight gain were not affected by dietary interventions. Proteomic analysis revealed 4 and 11 overabundant proteins in muscle and adipose tissues, respectively, in SOY compared to LIN. Additionally, the pro-inflammatory role of n-6 PUFAs may be associated to the observed overabundance of haptoglobin, an acute-phase protein, and the stimulation of protein-coding genes and proteins relevant to the innate immune response and acute inflammatory response. Using the same population, we applied the use of exon-intron split analysis (EISA) approach to account for post-transcriptional changes, driven by putative functional miRNA repression, in response to extreme values of n-6/n-3 PUFA ratio in longissimus dorsi muscle in male and female piglets. EISA revealed 11 and 97 mRNA genes within the top 5% negative PTc scores with at least 1.5-fold exonic region reduction in SOY-male (SOY-M) and LIN-male (LIN-M) piglets, respectively. Furthermore, the detected upregulated miRNAs harbored binding sites on 72.73% (SOY-M) and 61.86% (LIN-M) of these post-transcriptionally downregulated genes. Moreover, the overall PTc signals in females were not as strong and clear as those from the males. In addition, we discovered genes that are predominantly associated in the modulation of immune responses and lipid-related metabolism, which may be related to the pro- and anti-inflammatory properties of n-6 and n-3 PUFAs, respectively. Our use of EISA enabled us to identify regulatory networks that complemented traditional analyses of differential expression, providing us a more comprehensive understanding of muscle metabolic changes in response to PUFA concentration.