TY - JOUR
T1 - Enhanced ectopic bone formation using a combination of plasmid DNA impregnation into 3-D scaffold and bioreactor perfusion culture
AU - Hosseinkhani, Hossein
AU - Yamamoto, Masaya
AU - Inatsugu, Yasuyuki
AU - Hiraoka, Yosuke
AU - Inoue, Sachiko
AU - Shimokawa, Hitoyata
AU - Tabata, Yasuhiko
N1 - Funding Information:
The Japan Society for the Promotion of Science (JSPS) is greatly acknowledged for the support of this work. We gratefully acknowledge the technical assistance of Dr. Akihiro Kanematsu. We appreciate Faculty of Dentistry, Tokyo Medical and Dental University, Japan for providing a pBacBH2.
PY - 2006/3
Y1 - 2006/3
N2 - The objective of this study is to enhance in vivo ectopic bone formation by combination of plasmid DNA impregnation into three-dimensional (3-D) cell scaffolds and a developed in vitro culture method. Gelatin was cationized by introducing spermine (Sm) to the carboxyl groups for complexation with the plasmid DNA. As the MSC scaffold, collagen sponge reinforced by incorporation of poly(glycolic acid) (PGA) fibers was used. A complex of the cationized gelatin and plasmid DNA of BMP-2 was impregnated into the scaffold. MCS were seeded into each scaffold and cultured by a static and perfusion methods. When MSC were cultured in the PGA-reinforced collagen sponge, the level of BMP-2 expression was significantly enhanced by the perfusion culture compared with static method. When the osteoinduction activity of the PGA-reinforced collagen sponges seeded with PBS, MSC, naked plasmid DNA-BMP-2, cationized gelatin-plasmid DNA-BMP-2 complex, and transfected MSC by static and perfusion method, were studied following the implantation into the back subcutis of rats in terms of histological and biochemical examinations, homogeneous bone formation was histologically observed throughout the sponges seeded with cationized gelatin-plasmid DNA of BMP-2 complex and transfected MSC by perfusion method, although the extent of bone formation was higher for the later one. The level of alkaline phosphatase activity and osteocalcin content at the implanted sites of sponges seeded with transfected MSC by perfusion method were significantly high compared with those seeded with other agents. We conclude that combination of plasmid DNA-impregnated PGA-reinforced collagen sponge and the perfusion method was promising to promote the in vitro gene expression for MSC and in vivo ectopic bone formation.
AB - The objective of this study is to enhance in vivo ectopic bone formation by combination of plasmid DNA impregnation into three-dimensional (3-D) cell scaffolds and a developed in vitro culture method. Gelatin was cationized by introducing spermine (Sm) to the carboxyl groups for complexation with the plasmid DNA. As the MSC scaffold, collagen sponge reinforced by incorporation of poly(glycolic acid) (PGA) fibers was used. A complex of the cationized gelatin and plasmid DNA of BMP-2 was impregnated into the scaffold. MCS were seeded into each scaffold and cultured by a static and perfusion methods. When MSC were cultured in the PGA-reinforced collagen sponge, the level of BMP-2 expression was significantly enhanced by the perfusion culture compared with static method. When the osteoinduction activity of the PGA-reinforced collagen sponges seeded with PBS, MSC, naked plasmid DNA-BMP-2, cationized gelatin-plasmid DNA-BMP-2 complex, and transfected MSC by static and perfusion method, were studied following the implantation into the back subcutis of rats in terms of histological and biochemical examinations, homogeneous bone formation was histologically observed throughout the sponges seeded with cationized gelatin-plasmid DNA of BMP-2 complex and transfected MSC by perfusion method, although the extent of bone formation was higher for the later one. The level of alkaline phosphatase activity and osteocalcin content at the implanted sites of sponges seeded with transfected MSC by perfusion method were significantly high compared with those seeded with other agents. We conclude that combination of plasmid DNA-impregnated PGA-reinforced collagen sponge and the perfusion method was promising to promote the in vitro gene expression for MSC and in vivo ectopic bone formation.
KW - Cationization
KW - Enhanced gene expression
KW - In vitro culture system
KW - Osteoinduction
KW - PGA-reinforced collagen sponge
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U2 - 10.1016/j.biomaterials.2005.08.017
DO - 10.1016/j.biomaterials.2005.08.017
M3 - Article
C2 - 16139884
AN - SCOPUS:28444487378
VL - 27
SP - 1387
EP - 1398
JO - Biomaterials
JF - Biomaterials
SN - 0142-9612
IS - 8
ER -