title: Immobilization of FLAG-Tagged Recombinant Adeno-Associated Virus 2 onto Tissue Engineering Scaffolds for the Improvement of Transgene Delivery in Cell Transplants.
abstract: The technology of virus-based genetic modification in tissue engineering has provided the opportunity to produce more flexible and versatile biomaterials for transplantation. Localizing the transgene expression with increased efficiency is critical for tissue engineering as well as a challenge for virus-based gene delivery. In this study, we tagged the VP2 protein of type 2 adeno-associated virus (AAV) with a 3×FLAG plasmid at the N-terminus and packaged a FLAG-tagged recombinant AAV2 chimeric mutant. The mutant AAVs were immobilized onto the tissue engineering scaffolds with crosslinked anti-FLAG antibodies by N-succinimidyl-3-(2-pyridyldithiol) propionate (SPDP). Cultured cells were seeded to scaffolds to form 3D transplants, and then tested for viral transduction both in vitro and in vivo. The results showed that our FLAG-tagged AAV2 exerted similar transduction efficiency compared with the wild type AAV2 when infected cultured cells. Following immobilization onto the scaffolds of PLGA or gelatin sponge with anti-FLAG antibodies, the viral mediated transgene expression was significantly improved and more localized. Our data demonstrated that the mutation of AAV capsid targeted for antibody-based immobilization could be a practical approach for more efficient and precise transgene delivery. It was also suggested that the immobilization of AAV might have attractive potentials in applications of tissue engineering involving the targeted gene manipulation in 3D tissue cultures.
authors.affiliation: Department of Oral and Maxillofacial-Head and Neck Oncology, Beijing Stomatological Hospital, Capital Medical University, Beijing, China.
authors.forename: Hua
authors.initials: H
authors.lastname: Li
authors.affiliation: National Institutes for Food and Drug Controls, Beijing, China.
authors.forename: Feng-Lan
authors.initials: FL
authors.lastname: Zhang
authors.affiliation: Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing, China; Department of Medical Genetics, Capital Medical University, Beijing, China.
authors.forename: Wen-Jie
authors.initials: WJ
authors.lastname: Shi
authors.affiliation: Department of Medical Genetics, Capital Medical University, Beijing, China.
authors.forename: Xue-Jia
authors.initials: XJ
authors.lastname: Bai
authors.affiliation: Beijing Institute of Genomics, Chinese Academy of Sciences, Beijing, China.
authors.forename: Shu-Qin
authors.initials: SQ
authors.lastname: Jia
authors.affiliation: Department of Medical Genetics, Capital Medical University, Beijing, China.
authors.forename: Chen-Guang
authors.initials: CG
authors.lastname: Zhang
authors.affiliation: Department of Medical Genetics, Capital Medical University, Beijing, China; Beijing Institute of Brain Disorders, Beijing, China.
authors.forename: Wei
authors.initials: W
authors.lastname: Ding
mesh_terms: D041721:3T3-L1 Cells; D000818:Animals; D036022:Capsid Proteins / Q000235:genetics / Q000378:metabolism*; D017690:Cell Transplantation; D000229:Dependovirus / Q000235:genetics*; D016503:Drug Delivery Systems; D015316:Genetic Therapy; D005822:Genetic Vectors / Q000008:administration & dosage*; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D009842:Oligopeptides / Q000235:genetics / Q000378:metabolism*; D023822:Tissue Engineering; D054457:Tissue Scaffolds; D014161:Transduction, Genetic; D019076:Transgenes / Q000235:genetics*
chemical_list: D036022:Capsid Proteins; D009842:Oligopeptides; C442553:VP2 protein, Dependovirus; D049452:Green Fluorescent Proteins; C085706:FLAG peptide