Department of Anatomy, Institute of Biomedical Sciences, University of Sao Paulo, Sao Paulo 05508000, Brazil.
Faculty for Clinical Medicine Mannheim of the University of Heidelberg, Institute for Integrative Pathophysiology, Universitätsmedizin Mannheim, 68169 Mannheim, Germany.
Cells. 2020 Dec 2;9(12):2582. doi: 10.3390/cells9122582.
In this study we surveyed a rat skeletal muscle RNA-Seq for genes that are induced by hindlimb immobilization and, in turn, become attenuated by leucine supplementation. This approach, in search of leucine-atrophy protection mediating genes, identified histone deacetylase 4 () as highly responsive to both hindlimb immobilization and leucine supplementation. We then examined the impact of leucine on expression, tissue localization, and target genes. A total of 76 male Wistar rats (280 g) were submitted to hindlimb immobilization and/or leucine supplementation for 3, 7 and 12 days. These animals were euthanized, and soleus muscle was removed for further analysis. RNA-Seq analysis of hindlimb immobilized rats indicated a sharp induction (log2 = 3.4) of expression which was attenuated by leucine supplementation (50%). Real-time PCR and protein expression analysis by Western blot confirmed increased mRNA after 7 days of hindlimb immobilization and mitigation of induction by leucine supplementation. Regarding the localization, the proportion of positive nuclei was higher in the immobilized group and decreased after leucine supplementation. Also, we found a marked decrease of myogenin and - mRNA levels upon leucine supplementation, while and mRNA levels were increased by leucine supplementation. Our data suggest that inhibition might be involved in the anti-atrophic effects of leucine.
在这项研究中，我们调查了大鼠骨骼肌的 RNA-Seq，以寻找受下肢固定诱导、并转而被亮氨酸补充抑制的基因。这种方法旨在寻找亮氨酸抗萎缩保护介导基因，鉴定组蛋白去乙酰化酶 4 () 对下肢固定和亮氨酸补充均高度敏感。然后，我们研究了亮氨酸对表达、组织定位和靶基因的影响。总共 76 只雄性 Wistar 大鼠（280 克）接受下肢固定和/或亮氨酸补充治疗 3、7 和 12 天。这些动物被安乐死，取出比目鱼肌进行进一步分析。下肢固定大鼠的 RNA-Seq 分析表明表达显著诱导（log2 = 3.4），亮氨酸补充抑制诱导（50%）。实时 PCR 和 Western blot 蛋白表达分析证实，下肢固定 7 天后 mRNA 增加，亮氨酸补充减轻诱导。关于的定位，固定组阳性核的比例较高，亮氨酸补充后减少。此外，我们发现亮氨酸补充后肌生成素和 -mRNA 水平明显下降，而 - 和 -mRNA 水平则升高。我们的数据表明，抑制可能参与亮氨酸的抗萎缩作用。
