Composition comprising EPA, MA and leucine for improving muscle function

Abstract

The invention relates to compositions comprising EPA, MA and leucine for prevention and/or treatment of a disease or condition involving muscle decline or for improving muscle function.

Claims

1.-18. (canceled)

19. A method for improving muscle function of a subject, by administering to the subject a composition comprising eicosapentaenoic acid (20:5(n-3); EPA), myristic acid (C14:0; MA) and leucine, wherein the composition comprises EPA and MA in a weight ratio in the range of 1.0:1-1.6:1.

20. The method according to claim 19, wherein improving muscle function involves gaining muscle mass and/or maintaining muscle mass.

21. The method according to claim 19, wherein prevention and/or treatment of a disease or condition involving muscle decline and improving muscle function involve improving muscle protein synthesis.

22. The method according to claim 19, wherein the composition comprises less than 5 wt. % DHA based on fatty acids.

23. The method according to claim 19, wherein the composition comprises 0.1-1 g EPA per 100 ml of the composition.

24. The method according to claim 19, wherein at least 50 wt. % of the EPA and/or the MA is provided as free fatty acid.

25. The method according to claim 24, wherein all of the EPA and MA is provided as free fatty acid.

26. The method according to claim 19, wherein the composition comprises whey protein.

27. The method according to claim 19, wherein the composition comprises at least 11 wt. % leucine based on total proteinaceous matter.

28. The method according to claim 19, wherein the composition comprises at least 20 wt. % of leucine in a free form, relative to the total amount of leucine.

29. The method according to claim 19, wherein the composition further comprises one or more dietary fibres.

30. The method according to claim 19, wherein the composition further comprises carotenoids, vitamin A, vitamin B6, vitamin C, vitamin D3, vitamin E, folic acid, vitamin B12, selenium and/or zinc.

31. The method according to claim 19, wherein the composition is a supplement.

32. The method according to claim 19, for use in the treatment and/or prevention of sarcopenia, insufficient muscle protein synthesis, muscle degradation, muscle proteolysis, muscle atrophy, muscle dystrophy, muscle catabolism, muscle wasting, loss of muscle strength, loss of physical capacity and loss of physical performance.

33. The method according to claim 19, for the treatment and/or prevention of muscle decline and/or loss of muscle mass in a subject during or following body weight maintenance, during or following energy restriction, during or following bed rest or during recovery following physical trauma.

34. The method according to claim 19, wherein the subject is suffering from overweight or obesity, said subject following a weight loss program, an energy restriction program, a rehabilitation program and/or an exercise program.

Description

BRIEF DESCRIPTION OF THE FIGURES

[0093] FIG. 1 shows the results on puromycin incorporation of the in vitro study described in Example 1.

EXAMPLES

Example 1: In Vitro Experiment on Effects of EPA, MA and Leucine on Muscle Protein Synthesis

[0094] An experimental in vitro study is performed to evaluate the synergistic or additional action of EPA, MA and leucine on intracellular pathways regulating protein synthesis in muscle cells.

[0095] C2C12 mouse myoblasts are obtained from the American Type Culture Source Collection (no. CRL-1772). Myoblasts are cultured at 37° C. in an atmosphere of 5% CO2 in grown medium consisting of Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% fetal calf serum and antibiotics. Myotube C2C12 differentiation is induced by withdrawing fetal calf serum from confluent cells and adding 10 μg/ml insulin, 5 μg/ml transferrin and 2% horse serum.

[0096] 8-days-differentiated C2C12 myotubes are treated with medium containing EPA and/or MA for 16 hours. After a 4-hour starvation period, C2C12 myotubes were treated with insulin (100 nM)+Leucine (5 mM) for in total 60 min. A limitation of growing myotubes in vitro is that the cells are not able to produce insulin when administered leucine whereas in vivo the pancreas starts producing insulin when leucine is administered to a subject. Common to this model, to compensate for this limitation, the in vitro experiment requires the separate administration of insulin. Subsequently, a protein synthesis label (puromycin) was added to the medium and the tubes were homogenized in lysis buffer. By western-Blot analyses the level of protein synthesis was analyzed. This gives quantitative information on the incorporation of puromycin which is a measure for intracellular pathways regulating protein synthesis (Akt, mTOR) and/or a direct measure of protein synthesis. Hence the results are indicative for the prevention and/or treatment of a disease or condition involving muscle decline or for improving muscle function.

[0097] The model is widely applied in the art, to the same end i.e. drawing conclusions on muscle protein synthesis, muscle mass and muscle function. Reference is made to Brooks Mobley et al. “whey protein-derived exosomes increase protein synthesis and hypertrophy in C2C12 myotubes” J. Dairy Sci. 100:48-64 (2006); Jing et al. “alpha-lipoic acids promote the protein synthesis of C2C12 myotubes by the TLR2/PI13K signalling pathway” J. Agric. Food Chem. 2016, 64, 1720-1729; and Salles et al. “1,25(OH)2-vitamin D3 enhances the stimulating effect of leucine and insulin on protein synthesis rate through Akt/PKB and mTOR mediated pathways in murine C2C12 skeletal myotubes” Mol. Nutr. Food Res. 2013, 00, 1-10.

[0098] The above-described myoblasts were supplemented with various amounts of EPA and MA to assess their synergistic or additional action. The experimental legs all have muscle stimulation in the form of insulin and leucine. There is a control without stimulation and without fatty acids and there is a control with stimulation and without fatty acids. The experimental legs have: [0099] 25 μM of EPA without MA, [0100] 25 μM of MA without EPA, [0101] 50 μM of MA without EPA, [0102] 25 μM of EPA and 25 μM of MA (molar ratio 1:1 which is equivalent to a weight ratio of 1.3:1), [0103] 25 μM of EPA and 50 μM of MA (molar ratio of 1:2 which is equivalent to a weight ratio of 0.66:1).

[0104] The results are presented in the FIG. 1. All results are relative to the control without any stimulation and without fatty acids (the control is indexed at 100). As can be seen from the graph, both EPA and MA individually have an effect on puromycin incorporation. However the effect of puromycin incorporation of EPA and MA is much larger when added simultaneously compared to the effects of the individual fatty acids. This effect even surpasses the sum of the individual effects relative to the control. Hence EPA and MA act in a synergistic manner when added simultaneously.

[0105] Furthermore, it is shown that the ratio of EPA to MA is more important than the total level of fatty acids, as raising the level of from 25 μM MA to 50 μM MA had a much smaller effect than adding 25 μM of EPA to 25 μM of MA.

[0106] In addition, the bar showing the incorporation of puromycin incorporation for 25 μM EPA and 50 μM MA (i.e. molar ratio of 1:2 which corresponds to a weight ratio of 0.66:1) is significantly lower than the bar for 25 μM of each (which corresponds to a weight ratio of 1.3:1).