Farm Animal Genetic Resource Exploration and Innovation Key Laboratory of Sichuan Province, Chengdu 611130, China.
Int J Mol Sci. 2018 Sep 19;19(9):2837. doi: 10.3390/ijms19092837.
Guanidinoacetic acid (GAA), an amino acid derivative that is endogenous to animal tissues including muscle and nerve, has been reported to enhance muscular performance. MicroRNA (miRNA) is a post-transcriptional regulator that plays a key role in nutrient-mediated myogenesis. However, the effects of GAA on myogenic differentiation and skeletal muscle growth, and the potential regulatory mechanisms of miRNA in these processes have not been elucidated. In this study, we investigated the effects of GAA on proliferation, differentiation, and growth in C2C12 cells and mice. The results showed that GAA markedly inhibited the proliferation of myoblasts, along with the down-regulation of cyclin D1 () and cyclin dependent kinase 4 () mRNA expression, and the upregulation of cyclin dependent kinase inhibitor 1A () mRNA expression. We also demonstrated that GAA treatment stimulated myogenic differentiation 1 () and myogenin () mRNA expression, resulting in an increase in the myotube fusion rate. Meanwhile, GAA supplementation promoted myotube growth through increase in total myosin heavy chain (MyHC) protein level, myotubes thickness and gastrocnemius muscle cross-sectional area. Furthermore, small RNA sequencing revealed that a total of eight miRNAs, including miR-133a-3p and miR-1a-3p cluster, showed differential expression after GAA supplementation. To further study the function of miR-133a-3p and miR-1a-3p in GAA-induced skeletal muscle growth, we transfected miR-133a-3p and miR-1a-3p mimics into myotube, which also induced muscle growth. Through bioinformatics and a dual-luciferase reporter system, the target genes of miR-133a-3p and miR-1a-3p were determined. These two miRNAs were shown to modulate the Akt/mTOR/S6K signaling pathway by restraining target gene expression. Taken together, these findings suggest that GAA supplementation can promote myoblast differentiation and skeletal muscle growth through miR-133a-3p- and miR-1a-3p-induced activation of the AKT/mTOR/S6K signaling pathway.
胍基乙酸（GAA）是一种氨基酸衍生物，内源性存在于包括肌肉和神经在内的动物组织中，已被报道可增强肌肉性能。microRNA（miRNA）是一种转录后调控因子，在营养介导的肌发生中发挥关键作用。然而，GAA 对成肌分化和骨骼肌生长的影响，以及 miRNA 在这些过程中的潜在调节机制尚未阐明。在这项研究中，我们研究了 GAA 对 C2C12 细胞和小鼠增殖、分化和生长的影响。结果表明，GAA 显著抑制成肌细胞的增殖，同时下调细胞周期蛋白 D1（）和细胞周期蛋白依赖性激酶 4（）mRNA 表达，并上调细胞周期蛋白依赖性激酶抑制剂 1A（）mRNA 表达。我们还表明，GAA 处理刺激肌生成分化 1（）和肌生成素（）mRNA 表达，导致肌管融合率增加。同时，GAA 补充通过增加总肌球蛋白重链（MyHC）蛋白水平、肌管厚度和比目鱼肌横截面积来促进肌管生长。此外，小 RNA 测序显示，GAA 补充后共有 8 种 miRNA（包括 miR-133a-3p 和 miR-1a-3p 簇）表现出差异表达。为了进一步研究 miR-133a-3p 和 miR-1a-3p 在 GAA 诱导的骨骼肌生长中的功能，我们将 miR-133a-3p 和 miR-1a-3p 模拟物转染到肌管中，也诱导了肌肉生长。通过生物信息学和双荧光素酶报告系统，确定了 miR-133a-3p 和 miR-1a-3p 的靶基因。这两种 miRNA 通过抑制靶基因表达来调节 Akt/mTOR/S6K 信号通路。总之，这些发现表明，GAA 补充可通过 miR-133a-3p 和 miR-1a-3p 诱导的 Akt/mTOR/S6K 信号通路激活来促进成肌细胞分化和骨骼肌生长。
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