In vitro construction of tissue-engineered bone with bone morphogenetic protein-2-transfected rabbit bone marrow mesenchymal stem cells and hydroxyapatite nanocomposite.
OBJECTIVE
The aim of this study was to investigate the adenovirus-mediated human bone morphogenetic protein-2 (Ad-BMP-2) transfection of rabbit bone marrow mesenchymal stem cells (rBMSCs), which, together with hydroxyapatite nanocomposite (Nano-HA), were used to construct tissue-engineered bone in vitro.


METHODS
Ad-BMP-2 adenovirus vector was prepared with the Gateway technique and was transfected into rBMSCs. Sol-flocculation method was employed to prepare Nano-HA. Immunohistochemistry, reverse transcription-polymerase chain reaction, and Western blot assay were performed to detect the BMP-2 expression in transfected cells. At 48 h after transfection, transfected cells were inoculated into the Nano-HA scaffold. After 3 and 5 days, scanning electron microscopy was performed to observe adherence and growth of these cells. The cells in the scaffold were harvested after digestion, and Western blot assay was performed to detect the BMP-2.


RESULTS
After transfection, the mRNA and protein of hBMP-2 were expressed at a high level. Scanning electron microscopy indicated that these cells were evenly distributed in the scaffold, with favorable adherence. In addition, the cells collected from the scaffold had a high expression of BMP-2.


CONCLUSIONS
Adenovirus-mediated hBMP-2 transfection was successfully performed in rBMSCs, and these cells inoculated into the Nano-HA scaffold had a high expression of hBMP-2 in the scaffold. Thus, this technique is feasible to construct tissue-engineered bone in vitro.
Departments of Orthopedics, the First Affiliated Hospital of Gannan Medical University, Ganzhou, Jiangxi Province, China.
Chunlei
C
He
Cheng
C
He
Jianwen
J
Mo
Xiaona
X
Fan
Guanglin
G
Ji
Wuyang
W
Liu
Dongbao
D
Wu
Weimin
W
Zhu
Daping
D
Wang
Hui
H
Gao
D000818:Animals; D001846:Bone Development / Q000502:physiology*; D055396:Bone Morphogenetic Protein 2 / Q000235:genetics / Q000378:metabolism*; D018786:Bone Substitutes / Q000138:chemical synthesis; D002454:Cell Differentiation; D002478:Cells, Cultured; D017886:Durapatite / Q000737:chemistry; D004867:Equipment Design; D005818:Genetic Engineering / Q000379:methods; D006801:Humans; D008422:Materials Testing; D059630:Mesenchymal Stem Cells / Q000166:cytology* / Q000502:physiology*; D053758:Nanoparticles / Q000737:chemistry; D010006:Osteoblasts / Q000166:cytology* / Q000502:physiology*; D010012:Osteogenesis / Q000502:physiology; D011817:Rabbits; D023822:Tissue Engineering / Q000295:instrumentation*; D054457:Tissue Scaffolds; D014162:Transfection
C525718:BMP2 protein, human; D055396:Bone Morphogenetic Protein 2; D018786:Bone Substitutes; D017886:Durapatite