Thomson David M, Gordon Scott E
Human Performance Laboratory, 363 Ward Sports Medicine Building, East Carolina University, Greenville, NC 27858, USA.
J Physiol. 2006 Jul 1;574(Pt 1):291-305. doi: 10.1113/jphysiol.2006.107490. Epub 2006 Apr 20.
Impaired overload-induced protein synthesis and growth in aged fast-twitch skeletal muscle may result from diminished responsiveness of signalling intermediates controlling protein translation. Yet, potential age-related signalling decrements have never been examined in direct parallel with impaired overload-induced muscle growth in any model. To this end, we used Western blotting to examine the contents and phosphorylation states of mammalian target of rapamycin (mTOR) and its downstream translational signalling intermediates, 70 kDa ribosomal protein S6 kinase (S6k), ribosomal protein S6 (rpS6), eukaryotic elongation factor 2 (eEF2), and eukaryotic initiation factor 4E-binding protein 1 (4E-BP1), in conjunction with impaired growth in 1 week overloaded fast-twitch plantaris muscles (via unilateral gastrocnemius ablation) of old (O; 30 months) versus young adult (YA; 8 months) male Fischer344 x Brown Norway rats. The significantly (P <or= 0.05) diminished growth (assessed by total muscle protein content) in overloaded O muscles (5.6 +/- 1.7 versus 19.3 +/- 2.9% in YA) was accompanied by significant impairments in the phosphorylation states of mTOR (Ser2448), S6k (impaired at the mTOR-specific Thr389 residue but not at Thr421/Ser424), rpS6 (Ser235/236) and 4E-BP1 (gel shift), as well as deficits in total eEF2 accretion. Moreover, in overloaded muscles across both age groups, phospho-S6k at Thr389 (but not at Thr421/Ser424), 4E-BP1 phosphorylation status, and total eEF2 accretion were all positively correlated with percentage muscle hypertrophy, and negatively correlated with the phosphorylation (Thr172) of 5'-AMP-activated protein kinase (AMPK; which inhibits translational signalling and protein synthesis in young muscle at rest). As previously published by ourselves, AMPK was hyperphosphorylated in O versus YA muscles used in the current investigation. The present results provide solid evidence that impaired overload-induced growth in aged fast-twitch muscle may partly result from multiple-level decrements in signalling pathway(s) controlling protein translation, and also provide an initial indication that AMPK hyperactivation with age may potentially lie upstream of these decrements.
衰老的快肌骨骼肌中，过载诱导的蛋白质合成及生长受损可能是由于控制蛋白质翻译的信号中间体反应性降低所致。然而，在任何模型中，尚未有研究直接将潜在的年龄相关信号转导减少与过载诱导的肌肉生长受损进行平行比较。为此，我们采用蛋白质免疫印迹法，检测雷帕霉素靶蛋白（mTOR）及其下游翻译信号中间体——70 kDa核糖体蛋白S6激酶（S6k）、核糖体蛋白S6（rpS6）、真核生物延伸因子2（eEF2）和真核生物起始因子4E结合蛋白1（4E-BP1）的含量及磷酸化状态，同时检测老年（O组；30月龄）和年轻成年（YA组；8月龄）雄性Fischer344×Brown Norway大鼠1周过载快肌比目鱼肌（通过单侧腓肠肌切除）中生长受损的情况。过载的老年大鼠肌肉（O组）中生长显著（P≤0.05）受损（通过总肌肉蛋白含量评估，O组为5.6±1.7%，YA组为19.3±2.9%），同时mTOR（Ser2448）、S6k（在mTOR特异性的Thr389位点受损，但在Thr421/Ser424位点未受损）、rpS6（Ser235/236）和4E-BP1（凝胶迁移）的磷酸化状态显著受损，且eEF2总积累量也有缺陷。此外，在两个年龄组的过载肌肉中Thr389位点的磷酸化S6k（但Thr421/Ser424位点未磷酸化）、4E-BP1磷酸化状态及eEF2总积累量均与肌肉肥大百分比呈正相关，与5'-AMP激活的蛋白激酶（AMPK；在静息状态下抑制年轻肌肉中的翻译信号转导和蛋白质合成）的磷酸化（Thr172）呈负相关。正如我们之前发表的研究结果所示，在本研究中使用的老年大鼠与年轻成年大鼠的肌肉中，AMPK均发生了过度磷酸化。目前的研究结果提供了确凿证据，表明衰老的快肌中过载诱导的生长受损可能部分是由于控制蛋白质翻译的信号通路在多个水平上的减少所致，同时也初步表明随着年龄增长AMPK的过度激活可能潜在地位于这些减少的上游。
