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[Bone marrow stromal cells co-modified with BMP-2 and bFGF gene]
Yi Li1, Huai-qing Chen, Min Cheng
1Institute of Biomedical Engineering, West China Medical Center of Sichuan University, Chengdu 610041, China. yi_lee0103@yahoo.com.cn
Aim:
To establish modified bone marrow stromal cells(BMSCs) which can express BMP-2 and bFGF stably.
Methods:
BMP-2 and bFGF gene were amplified by RT-PCR, and then cloned into the expression vector pcDNA3.0. After being confirmed by DNA sequencing, pcDNA3.0-BMP-2 and pcDNA3.0-bFGF were co-transfected into rat BMSCs with Lipofectamine 2000 reagent. The expression of BMP-2 and bFGF gene in rat BMSCs was detected by RT-PCR, Western blot, immunohistochemical staining and ELISA.
Results:
BMP-2 and bFGF gene were cloned, and their sequences were identical with those in GenBank. The expression plasmids, pcDNA3.0-BMP-2 and pcDNA3.0-bFGF, were constructed and co-transfected into rMSCs successfully. RT-PCR showed the mass transcription of BMP-2 and bFGF mRNA in transfected BMSCs. Western blot, immunohistochemical staining and ELISA confirmed the expression of BMP-2 and bFGF genes in transfected cells and in the supernatant.
Conclusion:
We have constructed the optimal rat BMSCs which can be used in bone tissue engineering.
