A cell-based evaluation of a non-essential amino acid formulation as a non-bioactive control for activation and stimulation of muscle protein synthesis using ex vivo human serum
A cell-based evaluation of a non-essential amino acid formulation as a non-bioactive control for activation and stimulation of muscle protein synthesis using ex vivo human serum
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DOI：
10.1371/journal.pone.0220757
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发表时间：
2019-11-19
期刊：
影响因子：
3.7
通讯作者：
Carson, Brian P.
中科院分区：
文献类型：
--
作者：
Patel, Bijal;Pauk, Martina;Carson, Brian P.
本研究的目的是比较用含等氮非必需氨基酸（NEAA）或乳清蛋白水解物（WPH）的餐后离体人血清条件培养基处理骨骼肌细胞对刺激C2 C12骨骼肌细胞肌肉蛋白质合成（MPS）的效果。以及餐后（进食）60分钟摄入WPH或NEAA（0.33 g.kg（-1）体重）。用离体人血清（20%）条件培养基处理C2 C12肌管4小时。MPS信号传导的激活（mTOR、P70 S6 K和4 E-BP 1的磷酸化），随后分别通过蛋白质印迹和翻译表面传感技术（SUnSET）体外测定MPS。而与禁食血清相比，由WPH喂养的血清调节的培养基显著增加mTOR（Ser 2448）、P70 S6 K和4 E-BP 1磷酸化（p< 0.01，p< 0.05）。此外，与NEAA饲喂血清相比，WPH饲喂血清的条件培养基对蛋白质信号传导和MPS的影响显著增加（p< 0.01，p< 0.05）。因此，这些体外研究结果表明，使用等氮NEAA作为比较不同蛋白质对MPS活化的生物活性的有效对照。
PurposeThe purpose of this study was to compare the effect of treating skeletal muscle cells with media conditioned by postprandial ex vivo human serum fed with either isonitrogenous Non-Essential Amino Acid (NEAA) or a whey protein hydrolysate (WPH) on stimulating Muscle Protein Synthesis (MPS) in C2C12 skeletal muscle cells.MethodsBlood was taken from six young healthy males following overnight fast (fasted) and 60 min postprandial (fed) ingestion of either WPH or NEAA (0.33 g.kg(-1) Body Mass). C2C12 myotubes were treated with media conditioned by ex vivo human serum (20%) for 4 h. Activation of MPS signalling (phosphorylation of mTOR, P70S6K and 4E-BP1) were determined in vitro by Western Blot and subsequent MPS were determined in vitro by Western Blot and surface sensing of translation technique (SUnSET) techniques, respectively.ResultsMedia conditioned by NEAA fed serum had no effect on protein signalling or MPS compared to fasted, whereas media conditioned by WPH fed serum significantly increased mTOR (Ser2448), P70S6K and 4E-BP1 phosphorylation (p< 0.01, p< 0.05) compared to fasted serum. Furthermore, the effect of media conditioned by WPH fed serum on protein signalling and MPS was significantly increased (p< 0.01, p< 0.05) compared to NEAA fed serum.ConclusionIn summary, media conditioned by NEAA fed serum did not result in activation of MPS. Therefore, these in vitro findings suggest the use of isonitrogenous NEAA acts as an effective control for comparing bioactivity of different proteins on activation of MPS.