Effects of recombinant retroviral vector mediated human insulin like growth factor-1 gene transfection on skeletal muscle growth in rat.
BACKGROUND
This study transferred a recombinant gene encoding human insulin like growth factor-1 (hIGF-1) into modified primary skeletal myoblasts with a retroviral vector (pLgXSN) and determined whether the hIGF-1 promoted growth of skeletal muscle in rat.


METHODS
hIGF-1cDNA was amplified in vitro from normal human liver cells by using RT-PCR and cloned into plasmid vector pLgXSN. The recombinant vector pLghIGF-1SN and control vector pLgGFPSN were transfected into packaging cell PT67 and G418 was used to select positive colony. Myoblasts were infected with a high titre viral supernatant and transduction efficiency was evaluated as GFP expression. The expression of hIGF-1 mRNA in myoblasts was investigated by immunocytochemistry and RT-PCR. MTT assays detected the growth of myoblasts in vitro. Myoblasts transduced with pLghIGF-1SN were injected into hind limb muscles of 10 - 12 week male SD rats. Formed tissues were harvested 4 weeks later. Myocyte diameter, mean weight of hind limb and body were measured to evaluate the skeletal muscle growth.


RESULTS
Recombinant retroviral plasmid vector pLghIGF-1SN was constructed successfully. The titre of the packaged recombinant retrovirus was 1 x 10(6) cfu/ml. The transfection rate of PT67 cells reached 100% after G418 screening. hIGF-1 expression was positive in myoblast-IGF-1. The proliferation rate of myoblast-IGF-1 in vitro was higher than GFP-myoblast or myoblast (P < 0.05). The mean weights of hind limb and body of rats injected myoblast-IGF-1 were higher than those of the rats injected with myoblast-GFP or myoblast (P < 0.05). Myocyte diameter had a significant increase in IGF-1 group compared to GFP group and myoblast group (P < 0.05).


CONCLUSIONS
The transfection of the human IGF-1 gene mediated by a retroviral vector can promote the growth of skeletal muscle in rats. Genetically modified primary skeletal myoblasts provide a possibly effective approach to treat some skeletal muscle diseases.
Rong
Shu-ling
SL
Department of Cardiology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China.
Lu
Yong-xin
YX
Liao
Yu-hua
YH
Wang
Xiao-lin
XL
Guo
He-ping
HP
Chang
Chao
C
Gao
Yan-zhang
YZ
Mi
Shao-hua
SH
Wan
Jian-ping
JP
D000818:Animals; D002478:Cells, Cultured; D004274:DNA, Recombinant / Q000235:genetics; D005822:Genetic Vectors; D007334:Insulin-Like Growth Factor I / Q000235:genetics* / Q000502:physiology; D018482:Muscle, Skeletal / Q000254:growth & development*; D032446:Myoblasts / Q000502:physiology; D051381:Rats; D017207:Rats, Sprague-Dawley; D012190:Retroviridae / Q000235:genetics*; D014162:Transfection
D004274:DNA, Recombinant; D007334:Insulin-Like Growth Factor I