The Effect of EPA and DHA on Skeletal Muscle Physiology and Energy Balance

Eicosapentaenoic acid (EPA) and Docosahexaenoic acid (DHA) supplementation commonly suggested during pregnancy confers many health benefits on offspring and more recently improves fetal BAT development and induces myoblast trans-differentiation into adiopocytes through activating PPARs expression. Embryonic myoblast is a highly flexible cell showing great capacity to change its phenotype into adipocyte in response to EPA and DHA supplementation in many in vitro models. However, a little is known about the identity of derived adipocytes, whether, they change into brown or white adipocyte, and in vivo evidence is still missing. Given that the myogeneis process is initiated at the mid of gestation and fetal BAT development is formed at the late stage of pregnancy, we sought to investigate the effect of EPA and DHA supplementation on fetal muscle growth and energy regulation. To achieve our goal, the study was mapped in two patterns: three in vitro experiments, devoted to test the effect of combined and isolated EPA and DHA on C2C12 undergoing differentiation into white and brown adipocytes, and in vivo experiment devoted to investigate the effect of maternal ingestion of EPA and DHA enriched diet on muscle growth, BAT activity, lipid metabolism regulation in liver, and browning of sWAT. C2C12 cell have frequently been used as a representative model of myoblasts in preclinical trials. In vitro, Cells were induced to differentiate into white or brown adipocytes using hormonal cocktail in the absence (CON) or presence of (50 µm) EPA and (50 µm) DHA in combination or separately. The results suggested that EPA and DHA treatments potentiate the route of C2C12 trans-differentiation into white- like adipocytes with a greater potency of DHA in compared to EPA. The effect was mediated via inhibiting myogenesis and mitochondrial biogenesis processes and up-regulating the expression of WAT signature genes and impairing the aquistion of brown adipocyte phenotype. Changing genes profile was concurrent with increasing lipid droplets formation and impairing mitochondrial function. In vivo, mice were fed a diet containing (3.05%) fish oil (FA) enriched with EPA and DHA or diet devoid of fish oil (CON) throughout the entire period of gestation and lactation. The results showed partial inconsistency with in vitro trials where transient increase in myogenesis regulating genes and MHC4 without increasing muscle mass were observed in FA treated group. An increasing intramuscular fat infiltration as a result of stimulating the expression of adipogenesis regulating genes was predominant in FA treated group at day 1 and 21 post-parturition. Also, maternal EPA/ DHA intake induced up-regulation the expression of fatty acids catabolism regulating genes in liver. Stimulating BAT development and activity and browning of subcutaneous white adipose tissue were demonstrated in FA treated group in weaned mice. Taken together, EPA and DHA supplementation is associated with ectopic lipid accumulation in skeletal muscle but not on the expense of myoblasts, and it can be suggested as an excellent therapeutic option to combat childhood obesity as a result of their potential role in stimulating energy expenditure transcriptional program

Gene regulation networks in early phase of Duchenne muscular dystrophy

The aim of this study was to analyze previously published gene expression data of skeletal muscle biopsies of Duchenne muscular dystrophy (DMD) patients and controls (gene expression omnibus database, accession #GSE6011) using systems biology approaches. We applied an unsupervised method to discriminate patient and control populations, based on principal component analysis, using the gene expressions as units and patients as variables. The genes having the highest absolute scores in the discrimination between the groups, were then analyzed in terms of gene expression networks, on the basis of their mutual correlation in the two groups. The correlation network structures suggest two different modes of gene regulation in the two groups, reminiscent of important aspects of DMD pathogenesis

Signalling role of skeletal muscle during exercise

Abstract Upon acute exercise skeletal muscle is immediately and heavily recruited, while other organs appear to play only a minor role during exercise. These other organs show significant changes and improvements in function, although they are not directly targeted by exercise. These improvements suggest that skeletal muscle can communicate with other organs. In the past fifteen years it became clear that skeletal muscle produces and secretes a variety of signalling proteins that are able to interact and communicate with other organs. These signalling proteins are called myokines and are likely the link between exercising muscle and the rest of the body. The aim of the research presented this thesis is to study the signalling role of skeletal muscle during exercise and to gain further insight in the local molecular changes in skeletal muscle induced by exercise. In the first part of this thesis the focus was on the local changes induced in skeletal muscle by acute exercise and exercise training. The aim was to gain more insight in the molecular basis of exercise-induced changes in skeletal muscle. First we performed a microarray analysis on human muscle biopsies taken from endurance, resistance and combined exercise training interventions. We showed that despite a substantial overlap between the three exercise training types, each of the exercise training types had an unique gene expression print. The gene expression print found in combined exercise training lacked some specific oxidative and PPAR related components compared to the gene expression print found in endurance exercise training. For acute exercise microarray analysis was performed on muscle biopsies taken before and after an one-legged cycling intervention from resting and exercising skeletal muscle. Results showed that acute exercise induced large gene expression changes in active skeletal muscle. Furthermore, results showed that acute exercise also induced gene expression changes in resting skeletal muscle and that these changes were likely systemically induced via free fatty acids. In the second part of this thesis the focus was on the signalling role of skeletal muscle during exercise. Secretome analysis was performed on the microarrays of the muscle biopsies taken before and after the one-legged cycling intervention. This secretome analysis resulted in a list of putative myokines of which a selection was measured in the plasma. These plasma measurements showed that CCL2 (MCP-1) and CX3CL1 (Fractalkine) increased plasma levels during acute exercise. The findings of the one-legged cycling study furthermore showed that Angptl4 mRNA levels were higher in the resting leg compared to the exercising leg. Follow-up studies using cell culture and mice models revealed that Angptl4 levels were increased in the resting leg via free fatty acids that activated PPARs. In the exercising leg the increased Angptl4 levels were inhibited via AMPK activation. This resulted in an influx of triglyceride derived fatty acids in the exercising, but not in the resting skeletal muscle. In conclusion, we showed that exercise not only elicits molecular changes in active or trained skeletal muscle, but also in non-active organs such as resting skeletal muscle. Furthermore, we were able to identify several myokines produced by skeletal muscle during exercise, of which CCL2, CX3CL1 and Angptl4 were the most promising. CCL2, CX3CL1 and Angptl4 all increased plasma levels during acute exercise. It remains unclear what the systemic role is of these myokines. For Angptl4 we were able to provide more insight in the mechanism and local functioning during exercise. We concluded that Angptl4 is important in the substrate distribution during exercise. From the results presented this thesis we conclude that skeletal muscle has an important signalling role during exercise, but that it remains unclear how important this signalling role is systemically.</p