Changes in Skeletal Muscle and Body Weight on Sleeping Beauty Transposon-Mediated Transgenic Mice Overexpressing Pig mIGF-1.
Changes in Skeletal Muscle and Body Weight on Sleeping Beauty Transposon-Mediated Transgenic Mice Overexpressing Pig mIGF-1.
复制标题
睡美人转座子介导的过度表达猪 mIGF-1 的转基因小鼠骨骼肌和体重的变化
DOI：
10.1007/s10528-018-9848-7
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发表时间：
2018-08
影响因子：
2.4
通讯作者：
Song C
中科院分区：
文献类型：
--
作者：
Gao B;Wang W;Wu H;Chen C;Shen D;Wang S;Chen W;Zhang L;Chan S;Song C
胰岛素样生长因子（igf -1）是哺乳动物中一种重要的生长因子，但局部肌肉特异性胰岛素样生长因子1 （mIGF-1）对骨骼肌发育的作用鲜有报道。为了确定猪igf -1在体内对机体发育和肌肉沉积的影响，并探讨其分子机制，我们建立了转基因小鼠模型，为制备猪内源igf -1基因转基因猪提供实验数据。利用猪骨骼α-肌动蛋白基因的5′和3′调控区构建了骨骼肌特异性表达载体。表达盒两侧是睡美人转座子(SB)-末端重复序列。该重组载体在小鼠成肌细胞和猪胎儿成纤维细胞中具有较强的促进绿色荧光蛋白（EGFP）报告基因表达的能力，但在猪肾细胞中不具有特异性。α-肌动蛋白调控因子驱动的EGFP水平显著强于巨细胞病毒启动子驱动的EGFP水平。上述结果表明，克隆的α-肌动蛋白调节因子能够有效驱动外源基因在成肌细胞中的特异性表达，并成功构建了SB转座子介导的骨骼肌特异性表达载体。为了验证猪mIGF-1对骨骼肌生长的影响，通过原核微注射SB介导的mIGF-1骨骼表达载体和SB转座酶表达质粒，制备了转基因小鼠。创始小鼠和下一代F1小鼠的转基因阳性率分别为30%（54/180）和90.1%（64/71）。mIGF-1基因可以在骨骼肌中特异性表达。转基因小鼠的mRNA和蛋白水平分别是野生型小鼠的15倍和3.5倍。从8周龄开始，F1转基因小鼠的体重显著高于野生型小鼠。16周龄转基因雄性小鼠腓肠肌石蜡切片显示，与野生型小鼠相比，每单位肌纤维数量增加。mIGF-1在小鼠骨骼肌中的过表达可促进肌纤维肥大和肌肉生成，增加成年小鼠的平均体重。SB转座子介导的转基因小鼠模型具有较高的转基因效率。
Insulin-like growth factor (IGF-I) is an important growth factor in mammals, but the functions of the local muscle-specific isoform of insulin-like growth factor 1 (mIGF-1) to skeletal muscle development have rarely been reported. To determine the effect of pig mIGF-1 on body development and muscle deposition in vivo and to investigate the molecular mechanisms, the transgenic mouse model was generated which can also provide experimental data for making transgenic pigs with pig endogenous IGF1 gene. We constructed a skeletal muscle-specific expression vector using 5′- and 3′-regulatory regions of porcine skeletal α-actin gene. The expression cassette was flanked with Sleeping Beauty transposon (SB)-inverted terminal repeats. The recombinant vector could strongly drive enhanced green fluorescence protein (EGFP) reporter gene expression specifically in mouse myoblast cells and porcine fetal fibroblast cells, but not in porcine kidney cells. The EGFP level driven by α-actin regulators was significantly stronger than that driven by cytomegalovirus promoters. These results indicated that the cloned α-actin regulators could effectively drive specific expression of foreign genes in myoblasts, and the skeletal muscle-specific expression vector mediated with SB transposon was successfully constructed. To validate the effect of pig mIGF-1 on skeletal muscle growth, transgenic mice were generated by pronuclear microinjection of SB-mediated mIGF-1 skeletal expression vector and SB transposase-expressing plasmid. The transgene-positive rates of founder mice and the next-generation F1 mice were 30% (54/180) and 90.1% (64/71), respectively. The mIGF-1 gene could be expressed in skeletal muscle specifically. The levels of mRNA and protein in transgenic mice were 15 and 3.5 times higher, respectively, than in wild-type mice. The body weights of F1 transgenic mice were significantly heavier than wild-type mice from the age of 8 weeks onwards. The paraffin-embedded sections of gastrocnemius from 16-week-old transgenic male mice showed that the numbers of myofibers per unit were increased in comparison with those in the wild-type mice. mIGF-1 overexpression in mice skeletal muscle may promote myofibers hypertrophy and muscle production, and increased the average body weight of adult mice. Transgenic mice models can be generated by the mediation of SB transposon with high transgene efficiency.
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影响因子：
4.8
作者：
COLEMAN, ME;DEMAYO, F;SCHWARTZ, RJ
通讯作者：
SCHWARTZ, RJ
影响因子：
5.3
作者：
GUNNING, P;HARDEMAN, E;KEDES, L
通讯作者：
KEDES, L
影响因子：
3
作者：
Carlson DF;Geurts AM;Garbe JR;Park CW;Rangel-Filho A;O'Grady SM;Jacob HJ;Steer CJ;Largaespada DA;Fahrenkrug SC
通讯作者：
Fahrenkrug SC
影响因子：
29
作者：
Izumiya, Yasuhiro;Hopkins, Teresa;Walsh, Kenneth
通讯作者：
Walsh, Kenneth
影响因子：
12.4
作者：
Shavlakadze, T;White, J;Grounds, MD
通讯作者：
Grounds, MD