IGF 1 GENE TRANSFER TO ACCELERATE MUSCLE RECOVERY FOLLOWING DISUSE

该子项目是利用NIH/NCRR资助的中心赠款提供的资源的许多研究子项目之一。子项目和研究者（PI）可能从另一个NIH来源获得主要资金，因此可以在其他CRISP条目中表示。所列机构为中心机构，不一定为研究者机构。胰岛素样生长因子I（IGF-I）是一种有效的肌源性因子，已被证明在肌肉再生和肌肉肥大中发挥关键作用。病毒介导的IGF-I基因转移上调肌肉大小并增加肌肉力量。目前尚不清楚IGF-I诱导的肌肉肥大是否是由现有肌纤维和卫星细胞中蛋白质合成增加和/或蛋白水解率降低介导的。目的：研究胰岛素样生长因子-I（IGF-I）过表达对体内肌肉蛋白质合成速率的影响。方法：将携带IGF-1基因的重组腺相关病毒（rAAV-1）注射于C57 BL 6小鼠左后肢，观察其对IGF-1基因表达的影响。该构建体由编码IGF-I的完整大鼠IGF-I cDNA、肌球蛋白轻链（MLC）1/3启动子和增强子以及SV 40多聚腺苷酸化序列组成。向六只小鼠（年龄=3周龄）的一条后肢的前室的间隙中注射80 μ 1含有约10 10 rAAV颗粒的10%甘油/PBS，靶向趾长伸肌（EDL）。转染后三个月，体内混合肌肉蛋白质合成率进行了测量，使用掺入静脉注射的L-[13 C]-phe到EDL肌肉蛋白。使用气相色谱-燃烧-同位素比质谱法定量结构蛋白中的[13 C]-phe丰度。配对样本t检验用于转染和对侧后肢之间的比较。结果：rAAV-IGF 1转染导致肌肉湿重和横截面积显著增加（12.1 ± 8.5%）和（9.9 ± 9.1%）（p0.05）。与对侧对照肢体相比，rAAV-IGF 1注射肢体中的体内混合肌肉蛋白合成率高26.9 ± 15.6%（p0.05）。在rAAV-IGF 1注射的肢体中，平均EDL肌肉蛋白质合成速率为10.5 ± 2.8%/天，在对侧肢体中为8.2 ± 1.9%/天。结论：这些结果表明，肌肉特异性腺相关病毒过表达IGF-I增加了混合肌肉蛋白质的合成速率，这有助于肌肉肥大。进一步的研究将确定EDL的收缩特性，以及在卫星细胞和成熟肌纤维中激活的IGF-I信号通路的组分。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Insulin-like growth factor I (IGF-I) is a potent myogenic factor that has been shown to play a critical role in muscle regeneration and muscle hypertrophy. Viral-mediated gene transfer of IGF-I upregulates muscle size and increases muscle strength. It is not clear whether IGF-I-induced muscle hypertrophy is mediated by an increase in protein synthesis and/or a decrease in proteolysis rates in existing myofibers and satellite cells. PURPOSE: The objective of this study was to investigate the effect of IGF-I overexpression on in vivo muscle protein synthesis rate. METHODS: The left hindlimb of young adult C57BL6 mice was injected with a recombinant adeno-associated virus vector for IGF-I (rAAV-1). The construct consisted of the entire rat IGF-I cDNA which encodes for IGF-I, a Myosin Light Chain (MLC) 1/3 promoter and enhancer, and SV40 polyadenlyation sequence. Six mice (age=3 weeks) were injected with 80ul of 10% glycerol/PBS containing approximately 10 10 rAAV particles into the interstitial space of the anterior compartment of one hindlimb, targeting the extensor digitorum longus (EDL). Three months after transfection, in vivo mixed muscle protein synthesis rates were measured using the incorporation of intravenously administered L-[13C]-phe into EDL muscle proteins. [13C]-phe abundance in the structural proteins was quantitated using gas chromatography-combustion-isotope ratio mass spectrometry. Paired samples t-tests were used for comparisons between transfected and contralateral hindlimbs. RESULTS: rAAV-IGF1 transfection resulted in a significant increase (12.1+8.5%) in muscle wet weight and cross-sectional area (9.9+9.1%)(p0.05). In vivo mixed muscle protein synthesis rate was 26.9+15.6% higher in the rAAV-IGF1 injected limbs compared to the contralateral control limb (p0.05). Average EDL muscle protein synthesis rate was 10.5+2.8%/day in the rAAV-IGF1 injected limb and 8.2+1.9%/day in the contralateral limb. CONCLUSIONS: These results demonstrate that muscle specific adeno-associated virus overexpression of IGF-I increased mixed muscle protein synthesis rate and this contributed to muscle hypertrophy. Further research will determine EDL contractile properties, and the components of the IGF-I signaling pathway that are activated in satellite cells and mature myofibers.