Adeno-associated virus-mediated bone morphogenetic protein-7 gene transfer induces C2C12 cell differentiation into osteoblast lineage cells.
OBJECTIVE
To investigate the effects of bone morphogenetic protein-7 (BMP7)-expressing recombinant adeno-associated virus (AAV) vector on the differentiation of C2C12 cells.


METHODS
AAV-BMP7 was packaged by infecting the stable cell clone BHK-21 (integrated with recombinant AAV vector plasmid pSNAV-BMP7) with recombinant herpes simplex virus type 1, which expresses AAV-2 Rep and Cap and possesses AAV packaging functions. Following infection with AAV-BMP7 at multiplicities of infection of 1 x 10(5) vector genomes per cell and subsequent culture, C2C12 cells were assessed qualitatively for BMP7 production, alkaline phosphatase activity, osteocalcin production and Cbfal and MyoD expression.


RESULTS
C2C12 cells transduced with AAV-BMP7 could produce BMP7 protein until d 28. Alkaline phosphatase in the cultured C2C12 cell lysate was elevated. Secreted osteocalcin in the culture medium was detectable at d 12 and Cbfal mRNA expression level was upregulated, coinciding with downregulation of MyoD in a temporal manner.


CONCLUSIONS
The present in vitro study demonstrated that AAV-BMP7 could infect and efficiently convert C2C12 cells from myoblasts into osteoblast lineage cells.
Yang
Min
M
Department of Orthopedics, Peking University Third Hospital, Beijing 100083, China.
Ma
Qing-jun
QJ
Dang
Geng-ting
GT
Ma
Kang-tao
KT
Chen
Ping
P
Zhou
Chun-yan
CY
D000469:Alkaline Phosphatase / Q000378:metabolism; D000818:Animals; D055419:Bone Morphogenetic Protein 7; D019485:Bone Morphogenetic Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000502:physiology; D002454:Cell Differentiation; D002460:Cell Line; D019070:Cell Lineage; D050659:Core Binding Factor Alpha 1 Subunit / Q000235:genetics; D000229:Dependovirus / Q000235:genetics*; D004797:Enzyme-Linked Immunosorbent Assay; D015870:Gene Expression; D005822:Genetic Vectors / Q000235:genetics; D006801:Humans; D017570:MyoD Protein / Q000235:genetics; D032446:Myoblasts / Q000166:cytology* / Q000378:metabolism; D010006:Osteoblasts / Q000166:cytology* / Q000378:metabolism; D015675:Osteocalcin / Q000096:biosynthesis; D039904:Pluripotent Stem Cells / Q000166:cytology / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D013997:Time Factors; D014162:Transfection; D016212:Transforming Growth Factor beta / Q000096:biosynthesis / Q000235:genetics* / Q000502:physiology
C525819:BMP7 protein, human; D055419:Bone Morphogenetic Protein 7; D019485:Bone Morphogenetic Proteins; D050659:Core Binding Factor Alpha 1 Subunit; D017570:MyoD Protein; D012333:RNA, Messenger; D016212:Transforming Growth Factor beta; D015675:Osteocalcin; D000469:Alkaline Phosphatase