Objective

Branched-chain amino acids (BCAAs) have been reported to inhibit several types of muscle atrophy via the activation of the mechanistic target of rapamycin complex 1 (mTORC1). However, we previously found that BCAA did not activate mTORC1 in growth hormone (GH)-deficient spontaneous dwarf rats (SDRs), and that GH restored the stimulatory effect of BCAAs toward the mTORC1. The objective of this study was to determine whether GH or Insulin-like growth factor-I (IGF-I) stimulated the expression of L-type amino acid transporters (LATs) that delivered BCAAs, and whether LATs were involved in the mTORC1 activation.

Design

After the continuous administration of GH, cross-sectional areas (CSAs) of muscle fibers and LAT mRNA levels in the skeletal muscles of SDRs were compared to those from the SDRs that received normal saline. The effect of GH and IGF-I on LAT mRNA levels were determined in L6 and C2C12 myocytes. The effects of 2-aminobicyclo[2.2.1]heptane-2-carboxylic acid (BCH), a blocker for LATs, and LAT1 siRNA on mTORC1 activation and cell functions were examined in C2C12 cells.

Results

GH increased LAT1 and LAT4 mRNA levels in accordance with the increase in CSAs of muscle fibers in SDRs. IGF-I, and not GH, increased LAT1 mRNA levels in cultured L6 myocytes. IGF-I also increased LAT1 mRNA level in another myocyte line, C2C12. Furthermore, IGF-I reduced LAT3 and LAT4 mRNA levels in both cell lines. GH reduced LAT3 and LAT4 mRNA levels in L6 cells. BCH decreased basal C2C12 cell proliferation and reduced IGF-I-induced phosphorylation of 4E-BP1 and S6K, both of which are mTORC1 targets, but LAT1 siRNA did not affect the phosphorylation. This suggests that BCH may exert its effect via other pathway than LAT1.

Conclusions: IGF-I increased LAT1 mRNA level in myocytes. However, the role of LAT1 in IGF-I-induced mTORC1 activation and cell functions remains unclear.

中文翻译：

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生长激素和胰岛素样生长因子-I（IGF-I）调节自发性矮小大鼠以及L6和C2C12心肌细胞肌肉中L型氨基酸转运蛋白的表达。

目的

据报道，支链氨基酸（BCAAs）通过激活雷帕霉素复合物1（mTORC1）的机械靶标来抑制几种类型的肌肉萎缩。但是，我们以前发现BCAA不会在生长激素（GH）缺乏的自发性矮小大鼠（SDR）中激活mTORC1，并且GH恢复了BCAA对mTORC1的刺激作用。这项研究的目的是确定GH或胰岛素样生长因子I（IGF-1）是否刺激了递送BCAA的L型氨基酸转运蛋白（LAT）的表达，以及LAT是否参与mTORC1激活。

设计

连续施用GH后，将SDR骨骼肌的肌纤维横截面积（CSA）和LAT mRNA水平与接受生理盐水的SDR进行比较。在L6和C2C12肌细胞中确定了GH和IGF-I对LAT mRNA水平的影响。在C2C12细胞中检查了2-氨基双环[2.2.1]庚烷-2-羧酸（BCH），LAT的阻滞剂和LAT1 siRNA对mTORC1激活和细胞功能的影响。

结果

GH根据SDR中肌肉纤维CSA的增加而增加LAT1和LAT4 mRNA的水平。IGF-1（而不是GH）增加了培养的L6心肌细胞中LAT1 mRNA的水平。IGF-1也增加了另一种心肌细胞C2C12中LAT1 mRNA的水平。此外，IGF-1降低了两种细胞系中的LAT3和LAT4 mRNA水平。GH降低了L6细胞中LAT3和LAT4 mRNA的水平。BCH降低了基础C2C12细胞增殖并降低了IGF-I诱导的4E-BP1和S6K的磷酸化，这两个都是mTORC1靶标，但LAT1siRNA不影响磷酸化。这表明BCH可能通过LAT1以外的途径发挥作用。

结论：IGF-1增加了心肌细胞中LAT1 mRNA的水平。然而，LAT1在IGF-I诱导的mTORC1激活和细胞功能中的作用仍不清楚。