Muscle Cellular and Molecular Physiology Research Group (MCMPRG), Institute of Sport and Physical Activity Research (ISPAR), University of Bedfordshire, UK; School of Health and Social Care, Bournemouth University, UK.
J Steroid Biochem Mol Biol. 2013 Nov;138:152-61. doi: 10.1016/j.jsbmb.2013.05.005. Epub 2013 May 25.
We investigated the ability of testosterone (T) to restore differentiation in multiple population doubled (PD) murine myoblasts, previously shown to have a reduced differentiation in monolayer and bioengineered skeletal muscle cultures vs. their parental controls (CON) (Sharples et al., 2011, 2012 [7,26]). Cells were exposed to low serum conditions in the presence or absence of T (100nM)±PI3K inhibitor (LY294002) for 72h and 7 days (early and late muscle differentiation respectively). Morphological analyses were performed to determine myotube number, diameter (μm) and myonuclear accretion as indices of differentiation and myotube hypertrophy. Changes in gene expression for myogenin, mTOR and myostatin were also performed. Myotube diameter in CON and PD cells increased from 17.32±2.56μm to 21.02±1.89μm and 14.58±2.66μm to 18.29±3.08μm (P≤0.05) respectively after 72h of T exposure. The increase was comparable in both PD (+25%) and CON cells (+21%) suggesting a similar intrinsic ability to respond to exogenous T administration. T treatment also significantly increased myonuclear accretion (% of myotubes expressing 5+ nuclei) in both cell types after 7 days exposure (P≤0.05). Addition of PI3K inhibitor (LY294002) in the presence of T attenuated these effects in myotube morphology (in both cell types) suggesting a role for the PI3K pathway in T stimulated hypertrophy. Finally, PD myoblasts showed reduced responsiveness to T stimulated mRNA expression of mTOR vs. CON cells and T also reduced myostatin expression in PD myoblasts only. The present study demonstrates testosterone administration improves hypertrophy in myoblasts that basally display impaired differentiation and hypertrophic capacity vs. their parental controls, the action of testosterone in this model was mediated by PI3K/Akt pathway.
我们研究了睾酮（T）恢复多群体倍增（PD）鼠肌母细胞分化的能力，此前的研究表明，与亲本对照（CON）相比，这些细胞在单层和生物工程化的骨骼肌培养物中分化能力降低（Sharples 等人，2011 年，2012 年[7,26]）。细胞在低血清条件下暴露于 T（100nM）±PI3K 抑制剂（LY294002）下 72h 和 7 天（分别为早期和晚期肌肉分化）。进行形态分析以确定肌管数量、直径（μm）和肌核积累作为分化和肌管肥大的指标。还进行了肌生成蛋白、mTOR 和肌肉生长抑制素的基因表达变化。CON 和 PD 细胞的肌管直径分别从 17.32±2.56μm 增加到 21.02±1.89μm 和 14.58±2.66μm 增加到 18.29±3.08μm（P≤0.05），在 T 暴露 72h 后。在 PD（+25%）和 CON 细胞（+21%）中，这种增加是相似的，表明对外源性 T 给药的反应具有相似的内在能力。T 处理还显著增加了两种细胞类型在 7 天暴露后的核积累（表达 5+核的肌管百分比）（P≤0.05）。在 T 存在下添加 PI3K 抑制剂（LY294002）减弱了肌管形态的这些效应（在两种细胞类型中），表明 PI3K 途径在 T 刺激的肥大中起作用。最后，PD 肌母细胞对 T 刺激的 mTOR mRNA 表达的反应性低于 CON 细胞，T 还降低了 PD 肌母细胞中肌肉生长抑制素的表达。本研究表明，睾酮给药可改善基础分化和肥大能力低于亲本对照的肌母细胞的肥大，在该模型中，睾酮的作用是通过 PI3K/Akt 途径介导的。
