[Differentiation of adipose-derived stem cells induced by recombinant adenovirus's containing fibers derived from B-group serotype 35-mediated bone morphogenetic protein].
OBJECTIVE
To investigate the differentiation of the adipose-derived adult stem cell (ADASC) induced by the recombinant adenovirus's containing fibers derived from B-group serotype 35 (rAd5/F35)-mediated human bone morphogenetic protein 7 (hBMP-7) gene and to explore a new cell source for the bone tissue engineering.


METHODS
The hBMP-7 gene was amplified with the pcDNA1.1/AMP-hBMP-7 plasmid as a formwork. After the purification, the gene fragment was cloned into the pDC316 carrier for the recombination of the plasmid of pDC316-hBMP-7. The 293 cells were co-transfected by the skeleton plasmid of pBHG-fiber5/35 and the shuttle plasmid of pDC316-hBMP-7, and the recombinant plasmid of Ad5/F35-hBMP-7 was obtained; the recombinant plasmid of Ad5/ F35-enhancd green fluorescent protein(EGFP) was obtained by the similar method. The rat ADASCs were cultured and transfected by the Ad5/F35-hBMP-7 plasmid and the Ad5/F35-EGFP plasmid, respectively; the remaining untransfected ADASC were used as the controls. The morphology and the growth pattern of the transfected cells were evaluated. The transcription and the expression of the transfected genes and the steogenic phenotypes such as calcium nodules and osteocalcin were evaluated by ELISA.


RESULTS
The identification of PCR and enzyme cutting showed that the construction of the recombinant Ad5/F35-hBMP-7 plasmid could be confirmed. The transfection rate of the ADASC by the Ad5/F35-EGFP plasmid was determined to be greater than 90%. The hBMP-7 gene in the transfected ADASC could express the corresponding protein, and the formation of the calcium nodules could be found in the induced group. The electron microscopy showed that there was a calcium element in the cytoplasm, the alkaline phosphatase result was positive, and the expression of osteocalcin was increased.


CONCLUSIONS
The rAd5/F35-hBMP-7 gene can promote the differentiation of the adipose-derived adult stem cells to the osteoblasts in the bone tissue engineering.
Kang
Yan
Y
Department of Orthopedics, First Affiliated Hospital of SUN Yet-Sen University, Guangzhou Guangdong , 510080, P. R. China. neokang@163.com
Liao
Weiming
W
Yuan
Zhenhua
Z
D000256:Adenoviridae / Q000235:genetics; D000273:Adipose Tissue / Q000166:cytology*; D000818:Animals; D055419:Bone Morphogenetic Protein 7 / Q000235:genetics* / Q000378:metabolism; D002454:Cell Differentiation; D002460:Cell Line; D015870:Gene Expression; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors / Q000235:genetics; D006801:Humans; D010006:Osteoblasts / Q000166:cytology* / Q000378:metabolism; D010957:Plasmids / Q000235:genetics; D051381:Rats; D011995:Recombination, Genetic; D013234:Stem Cells / Q000166:cytology*; D023822:Tissue Engineering; D014162:Transfection
D055419:Bone Morphogenetic Protein 7