title: Osteogenic differentiation of adipose-derived stem cells and calvarial defect repair using baculovirus-mediated co-expression of BMP-2 and miR-148b.
abstract: Repair of large calvarial bony defect remains a challenge for orthopedic surgeons. Since microRNAs (miRNAs) modulate the osteogenesis of osteoprogenitor cells, we aimed to engineer human adipose-derived stem cells (hASCs), a promising cell source for bone engineering, with miRNA-expressing baculovirus vectors. We constructed 4 baculoviruses each expressing 1 human miRNA (miR-26a, miR-29b, miR-148b, miR-196a) and verified that the miRNA-expressing baculovirus vectors augmented hASCs osteogenesis. Among these 4 miRNAs, miR-148b and miR-196a exerted more potent osteoinductive effects than miR-26a and miR-29b. Furthermore, we unveiled that co-transduction of hASCs with miR-148b-expressing and bone morphogenetic protein 2 (BMP-2)-expressing baculovirus vectors enhanced and prolonged BMP-2 expression, and synergistically promoted the in vitro osteogenic differentiation of hASCs. Implantation of the hASCs co-expressing BMP-2/miR-148b into critical-size (4 mm in diameter) calvarial bone defects in nude mice accelerated and potentiated the bone healing and remodeling, filling ≈94% of defect area and ≈89% of defect volume with native calvaria-like flat bone in 12 weeks, as judged from micro computed tomography, histology and immunohistochemical staining. Altogether, this study confirmed the feasibility of combining miRNA and growth factor expression for synergistic stimulation of in vitro osteogenesis and in vivo calvarial bone healing.
authors.affiliation: Department of Chemical Engineering, National Tsing Hua University, Hsinchu 300, Taiwan.
authors.forename: Ya-Hsin
authors.initials: YH
authors.lastname: Liao
authors.affiliation: College of Medicine, Chang Gung University, Taoyuan 333, Taiwan; Center for Tissue Engineering, Chang Gung Memorial Hospital, Taoyuan 333, Taiwan.
authors.forename: Yu-Han
authors.initials: YH
authors.lastname: Chang
authors.affiliation: Department of Chemical Engineering, National Tsing Hua University, Hsinchu 300, Taiwan.
authors.forename: Li-Yu
authors.initials: LY
authors.lastname: Sung
authors.affiliation: Department of Chemical Engineering, National Tsing Hua University, Hsinchu 300, Taiwan.
authors.forename: Kuei-Chang
authors.initials: KC
authors.lastname: Li
authors.affiliation: Department of Chemical Engineering, National Tsing Hua University, Hsinchu 300, Taiwan.
authors.forename: Chia-Lin
authors.initials: CL
authors.lastname: Yeh
authors.affiliation: College of Medicine, Chang Gung University, Taoyuan 333, Taiwan; Department of Nuclear Medicine and Molecular Imaging Center, Chang Gung Memorial Hospital, Taoyuan 333, Taiwan.
authors.forename: Tzu-Chen
authors.initials: TC
authors.lastname: Yen
authors.affiliation: Bioresource Collection and Research Center, Food Industry Research and Development Institute, Hsinchu 300, Taiwan.
authors.forename: Shiaw-Min
authors.initials: SM
authors.lastname: Hwang
authors.affiliation: Department of Nuclear Medicine and Molecular Imaging Center, Chang Gung Memorial Hospital, Taoyuan 333, Taiwan; Department of Medical Imaging and Radiological Sciences, Chang Gung University, Taoyuan 333, Taiwan.
authors.forename: Kun-Ju
authors.initials: KJ
authors.lastname: Lin
authors.affiliation: Department of Chemical Engineering, National Tsing Hua University, Hsinchu 300, Taiwan. Electronic address: ychu@mx.nthu.edu.tw.
authors.forename: Yu-Chen
authors.initials: YC
authors.lastname: Hu
mesh_terms: D000273:Adipose Tissue / Q000166:cytology; D000818:Animals; D016367:Baculoviridae / Q000235:genetics; D055396:Bone Morphogenetic Protein 2 / Q000235:genetics / Q000378:metabolism*; D002454:Cell Differentiation; D002478:Cells, Cultured; D005260:Female; D015870:Gene Expression; D005786:Gene Expression Regulation; D005822:Genetic Vectors / Q000235:genetics; D006801:Humans; D007150:Immunohistochemistry; D051379:Mice; D008807:Mice, Inbred BALB C; D008819:Mice, Nude; D035683:MicroRNAs / Q000235:genetics / Q000378:metabolism*; D010012:Osteogenesis; D012886:Skull / Q000293:injuries*; D013234:Stem Cells / Q000166:cytology*; D023822:Tissue Engineering; D054457:Tissue Scaffolds; D014945:Wound Healing; D055114:X-Ray Microtomography
chemical_list: C525718:BMP2 protein, human; D055396:Bone Morphogenetic Protein 2; C550570:MIRN148 microRNA, human; D035683:MicroRNAs
keywords: Adipose-derived stem cells; BMP-2; Baculovirus; Calvarial bone defect; Gene therapy; miR-148b