Date Published: October 4, 2018
Publisher: Public Library of Science
Author(s): Marika Vitali, Corrado Dimauro, Rubina Sirri, Martina Zappaterra, Paolo Zambonelli, Elisabetta Manca, Dalal Sami, Domenico Pietro Lo Fiego, Roberta Davoli, Juan J. Loor.
Porcine fat traits depend mostly on the interaction between nutritional and genetic factors. However, the pathways and biological processes influenced by this interaction are still poorly known in pigs, although they can have a huge impact on meat quality traits. The present research provides new knowledge insight into the effect of four diets (D1 = standard diet; D2 = linseed supplementation; D3 = linseed, vitamin E and selenium supplementation; D4 = linseed and plant-derived polyphenols supplementation) on the expression of 24 candidate genes selected for their role in lipid and energy metabolism. The data indicated that 10 out of 24 genes were differentially expressed among diets, namely ACACA, ADIPOQ, ADIPOR1, CHREBP (MLXPL), ELOVL6, FASN, G6PD, PLIN2, RXRA and SCD. Results from the univariate analysis displayed an increased expression of ACACA, ADIPOQ, ADIPOR1, CHREBP, ELOVL6, FASN, PLIN2, RXRA and SCD in D4 compared to D2. Similarly, ACACA, ADIPOQ, ADIPOR1, ELOVL6 and SCD were highly expressed in D4 compared to D3, while no differences were observed in D2-D3 comparison. Moreover, an increased expression of G6PD and ELOVL6 genes in D4 compared to D1 was observed. Results from the multivariate analysis confirmed that D2 was not different from D3 and that ACACA, SCD and FASN expression made D4 different from D2 and D3. Comparing D4 and D1, the expression levels of ELOVL6 and ACACA were the most influenced. This research provides evidence that the addition of both n-3 PUFA and polyphenols, derived from linseed, grape-skin and oregano supplementation in the diets, stimulates the expression of genes involved in lipogenesis and in oxidative processes. Results evidenced a greater effect on gene expression of the diet added with both plant extracts and n-3 PUFA, resulting in an increased expression of genes coding for fatty acid synthesis, desaturation and elongation in pig Longissimus thoracis muscle.
The phenotypic variability of porcine fat traits is regulated by many environmental and genetic factors, and several studies have demonstrated that both diet and genotype are the main factors influencing intramuscular fatty acid (FA) composition in all the animal species [1–3]. However, as the influence of diet on biological processes and pathways is poorly known [4–6], identification of genes involved in lipid metabolism and their relationship with diet is of main interest for meat quality research purposes in pigs. Many studies reported the importance of identifying the effect of lipogenic genes expression, in order to improve the knowledge on biological processes and metabolic pathways influencing muscle fat deposition to provide new insight into carcass adiposity [7,8]. In literature, several studies dealt with the effects of n-3 and n-6 polyunsaturated FA (PUFA) diet supplementation on lipid metabolism in skeletal muscle, adipose and liver tissues. These researches focused on the effect that different diets may have on the expression of genes involved in PUFA synthesis or in the regulation of FA composition [9–14]. However, the effect of PUFA dietary supplementation on the regulation of lipid metabolism through the modification of gene expression in swine is still poorly understood. De Tonnac et al.  found that the intake of docosahexaenoic acid (DHA) down-regulated the transcription of genes involved in fatty acid (FA) metabolism regulation such as FADS2 and SREBP1 in the liver and DECR2 in the Longissimus thoracis muscle of growing-finishing cross-breed pigs. In a study on the transcription profile of porcine Gluteus medius muscle, Ogłuszka et al. , showed that a diet supplemented with n-3 and n-6 PUFA led to the down-regulation of genes coding for apolipoproteins. Tous et al.  reported that dietary conjugated linoleic acid (CLA) can affect the expression of both porcine lipogenic and regulatory genes including PPARA, PPARG, FASN, SREBF1, ACACA, LPL, D6D, SCD in a tissue-specific manner. Some studies have described that supplementing the diet with antioxidants or polyphenols can also influence nutrient digestibility, gut microbiota, expression of pro-inflammatory genes and meat quality traits in pigs [15–19]. Overall, the effects of dietary supplementation on gene expression are mostly unknown. In particular, the effect of a diet enriched with both n-3 PUFA and antioxidants or polyphenols on the expression of genes involved in lipid metabolism of pig muscle has not been studied by other Authors until now.
In the present study, both univariate and multivariate analysis was applied in order to investigate the influence of four different diets on the expression level of a set of genes in porcine muscle. Genes were selected because of their role in lipid and energy metabolism and in the fat deposition of muscle tissue. Besides univariate statistics, which considers the effect of a single gene expression, multivariate analysis was conducted to account for the relationship (covariance and correlation) among the expressed genes, making the two approaches, complementary. Indeed, since the transcription levels of genes involved in the same metabolic pathways are often highly correlated, there is a growing trend to analyze gene expression data utilizing both univariate and multivariate approaches [27, 28].
The results obtained in the present study are useful to increase knowledge about the effects determined by experimental diets enriched in PUFA and antioxidants on the expression of skeletal muscle genes in pigs. Overall the dietary treatments produced effects on the expression of the tested genes. The use of both multivariate CDA and univariate ANOVA analyses allowed to better identify the gene interactions related to biological processes. Indeed, the results suggested that adding plant extract (source of polyphenols) and linseed (source of n-3 PUFA) stimulates the expression of genes involved in the control of muscle metabolism, leading to a mutual interaction between lipogenesis and oxidative processes in the Longissimus thoracis muscle of pigs. This stimulation was more evident in the diet supplemented with polyphenols and n-3 PUFA compared to a diet only enriched in n-3 PUFA. On the contrary, the integration of vitamin E and Selenium in the diet did not significantly alter the expression of the tested genes in comparison to the diet with only linseed supplementation. These results were a first step in the formulation of functional diets for pigs addressing consumers’ demand for healthy meat products.