College of Animal Science and Technology, Qingdao Agricultural University, Qingdao 266109, China; College of Animal Science, Inner Mongolia Key Laboratory of Equine Genetics, Breeding and Reproduction, Equine Research Center, Inner Mongolia Agricultural University, Hohhot 010018, China.
College of Animal Science, Inner Mongolia Key Laboratory of Equine Genetics, Breeding and Reproduction, Equine Research Center, Inner Mongolia Agricultural University, Hohhot 010018, China.
Comp Biochem Physiol Part D Genomics Proteomics. 2024 Sep;51:101249. doi: 10.1016/j.cbd.2024.101249. Epub 2024 May 20.
Previous research has shown that leucine (Leu) can stimulate and enhance the proliferation of equine skeletal muscle satellite cells (SCs). The gene expression profile associated with Leu-induced proliferation of equine SCs has also been documented. However, the specific role of Leu in regulating the expression of slow-twitch muscle fibers (slow-MyHC) and mitochondrial function in equine SCs, as well as the underlying mechanism, remains unclear. During this investigation, equine SCs underwent culturing in differentiation medium and were subjected to varying concentrations of Leu (0 mM, 0.5 mM, 1 mM, 2 mM, 5 mM, and 10 mM) over a span of 3 days. AMP-activated protein kinase (AMPK) inhibitor Compound C and mammalian target of rapamycin complex (mTOR) inhibitor Rapamycin were utilized to explore its underlying mechanism. Here we showed that the expression of slow-MyHC at 2 mM Leu level was significantly higher than the concentration levels of 0 mM,0.5 mM and 10 mM (P <0.01), and there was no significant difference compared to other groups (P > 0.05); the basal respiration, maximum respiration, standby respiration and the expression of slow-MyHC, PGC-1α, Cytc, ND1, TFAM, and COX1 were significantly increased with Leu supplementation (P < 0.01). We also found that Leu up-regulated the expression of key proteins on AMPK and mTOR signaling pathways, including LKB1, p-LKB1, AMPK, p-AMPK, S6, p-S6, 4EBP1, p-4EBP1, mTOR and p-mTOR (P < 0.05 or P < 0.01). Notably, when we treated the equine SCs with the AMPK inhibitor Compound C and the mTOR inhibitor Rapamycin, we observed a reduction in the beneficial effects of Leu on the expression of genes related to slow-MyHC and signaling pathway-related gene expressions. This study provides novel evidence that Leu promotes slow-MyHC expression and enhances mitochondrial function in equine SCs through the AMPK/mTOR signaling pathways, shedding light on the underlying mechanisms involved in these processes for the first time.
先前的研究表明亮氨酸（Leu）可以刺激和增强马骨骼肌卫星细胞（SCs）的增殖。Leu 诱导马 SC 增殖相关的基因表达谱也已被记录。然而，亮氨酸在调节马 SC 中慢肌纤维（slow-MyHC）和线粒体功能的表达中的具体作用以及潜在机制尚不清楚。在这项研究中，马 SC 在分化培养基中培养，并在 3 天内接受不同浓度的亮氨酸（0 mM、0.5 mM、1 mM、2 mM、5 mM 和 10 mM）处理。利用 AMP 激活的蛋白激酶（AMPK）抑制剂 Compound C 和雷帕霉素靶蛋白复合物（mTOR）抑制剂 Rapamycin 来探索其潜在机制。在这里，我们发现 2 mM Leu 水平下 slow-MyHC 的表达明显高于 0 mM、0.5 mM 和 10 mM 的浓度水平（P <0.01），与其他组相比没有显着差异（P > 0.05）；基础呼吸、最大呼吸、备用呼吸以及 slow-MyHC、PGC-1α、Cytc、ND1、TFAM 和 COX1 的表达均随 Leu 补充而显着增加（P <0.01）。我们还发现亮氨酸上调了 AMPK 和 mTOR 信号通路关键蛋白的表达，包括 LKB1、p-LKB1、AMPK、p-AMPK、S6、p-S6、4EBP1、p-4EBP1、mTOR 和 p-mTOR（P <0.05 或 P <0.01）。值得注意的是，当我们用 AMPK 抑制剂 Compound C 和 mTOR 抑制剂 Rapamycin 处理马 SC 时，我们观察到 Leu 对与 slow-MyHC 相关的基因和信号通路相关基因表达的有益作用降低。这项研究提供了新的证据，表明亮氨酸通过 AMPK/mTOR 信号通路促进马 SC 中 slow-MyHC 的表达并增强线粒体功能，首次揭示了这些过程涉及的潜在机制。
