CEN Shiqiang 1 , ZHANG Junmei 2 , HUANG Fuguo 1 , YANG Zhiming 3 , XIE Huiqi 3
  • 1Department of Orthopaedic s, West China Hospital , Sichuan University，Chengdu Sichuan, 610041，P. R. China;;
  • 2Cl inical Laboratory, the Third People’s Hospital of Chengdu;;
  • 3Division of Stem Cell and Tissue Engineering, State Key Laboratory of Biotherapy, West China Hospital , Sichuan University.;
Tissue engineering; Myoblast; IGF-1; Flow cytometry; In vitro culture
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【Abstract】 Objective To investigate the effect of IGF-1 on the growth of primary human embryonic
myoblasts. Methods The method of incorporation of 3H-TdR was used to evaluate the abil ity of prol iferation of myoblasts.The count per minute (CPM) values of myoblasts at different concentrations(1, 2, 4, 8, 16 and 32 ng/mL) of IGF-1 were measured,and dose-effect curves were drawn to choose the optional concentration of IGF-1 to promote the prol iferation. Then theexperimental group of myoblasts received the addition of the optional concentration of IGF-1 in the growth medium, the controlgroup just received the growth medium. The flow cytometry was used to detect the cell cycle . The method of incorporation of3H-TdR was used to measure the peak-CPM. The myotube fusion rate was measured in myoblasts with different concentrations(0, 5,
10, 15, 20, 25 and 30 ng/ mL) of IGF-1 in fusion medium, the dose-effect curves were also drawn, so as to decided the optional concentrationof IGF-1 in stimulating differentiation. Fusion medium with optional concentration of IGF-1 was used in experimentalgroup, and the control group just with fusion medium. The fusion rate of myotube and the synthesis of creatine kinase(CK) weredetected in both groups. Results The optional concentration of 5 ng/mL IGF-1 was chosen for stimulating prol iferation . It was shown that the time of cell cycle of control was 96 hours, but that of the experimental group was reduced to 60 hours. The results of flow cytometry showed that the time of G1 phase, S phase and G2M phase was 70.03, 25.01 and 0.96 hoursrespectively in control group, and were 22.66, 16.47 and 20.87 hours respectively in experimental group. The time-CPM value curves showed that the peak-CPM emerged at 96 hours in control group and 48 hours in experimental group, which was in agreement
with the results of the flow cytometry. The optional concentration stimulating prol iferation was 20 ng/mL IGF-1. Compared with control, the quantity of CK was increased by 2 000 mU/mL and the fusion rate was elevated by 30% in experimental group. Conclusion The concentrations of 20 ng/mL IGF-1 can elevat obviously the fusion rate and the quantity of CK. IGF-1 can enhance the prol iferation and differentiation of myoblasts via inducing the number of myoblasts at G1 phase and increasing the number of myoblasts at S and G2M phases.

Citation： CEN Shiqiang,ZHANG Junmei,HUANG Fuguo,YANG Zhiming,XIE Huiqi. EFFECT OF IGF-1 ON PROLIFERATION AND DIFFERENTIATION OF PRIMARY HUMAN EMBRYONIC MYOBLASTS. Chinese Journal of Reparative and Reconstructive Surgery, 2008, 22(1): 84-87. doi: Copy