title: Gene delivery to cartilage defects using coagulated bone marrow aspirate.
abstract: The long-term goal of the present study is to develop a clinically applicable approach to enhance natural repair mechanisms within cartilage lesions by targeting bone marrow-derived cells for genetic modification. To determine if bone marrow-derived cells infiltrating osteochondral defects could be transduced in situ, we implanted collagen-glycosaminoglycan (CG) matrices preloaded with adenoviral vectors containing various marker genes into lesions surgically generated in rabbit femoral condyles. Analysis of the recovered implants showed transgenic expression up to 21 days; however, a considerable portion was found in the synovial lining, indicating leakage of the vector and/or transduced cells from the matrix. As an alternative medium for gene delivery, we investigated the feasibility of using coagulated bone marrow aspirates. Mixture of an adenoviral suspension with the fluid phase of freshly aspirated bone marrow resulted in uniform dispersion of the vector throughout, and levels of transgenic expression in direct proportion to the density of nucleated cells in the ensuing clot. Furthermore, cultures of mesenchymal progenitor cells, previously transduced ex vivo with recombinant adenovirus, were readily incorporated into the coagulate when mixed with fresh aspirate. These vector-seeded and cell-seeded bone marrow clots were found to maintain their structural integrity following extensive culture and maintained transgenic expression in this manner for several weeks. When used in place of the CG matrix as a gene delivery vehicle in vivo, genetically modified bone marrow clots were able to generate similarly high levels of transgenic expression in osteochondral defects with better containment of the vector within the defect. Our results suggest that coagulates formed from aspirated bone marrow may be useful as a means of gene delivery to cartilage and perhaps other musculoskeletal tissues. Cells within the fluid can be readily modified with an adenoviral vector, and the matrix formed from the clot is completely natural, native to the host and is the fundamental platform on which healing and repair of mesenchymal tissues is based.
authors.affiliation: Center for Molecular Orthopaedics, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02115, USA.
authors.forename: A
authors.initials: A
authors.lastname: Pascher
authors.forename: G D
authors.initials: GD
authors.lastname: Palmer
authors.forename: A
authors.initials: A
authors.lastname: Steinert
authors.forename: T
authors.initials: T
authors.lastname: Oligino
authors.forename: E
authors.initials: E
authors.lastname: Gouze
authors.forename: J-N
authors.initials: JN
authors.lastname: Gouze
authors.forename: O
authors.initials: O
authors.lastname: Betz
authors.forename: M
authors.initials: M
authors.lastname: Spector
authors.forename: P D
authors.initials: PD
authors.lastname: Robbins
authors.forename: C H
authors.initials: CH
authors.lastname: Evans
authors.forename: S C
authors.initials: SC
authors.lastname: Ghivizzani
mesh_terms: D000256:Adenoviridae / Q000235:genetics*; D000818:Animals; D016026:Bone Marrow Transplantation; D002357:Cartilage Diseases / Q000473:pathology / Q000628:therapy*; D015870:Gene Expression; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors / Q000008:administration & dosage*; D023421:Models, Animal; D011817:Rabbits; D033581:Stem Cell Transplantation; D014161:Transduction, Genetic / Q000379:methods; D019076:Transgenes; D014182:Transplantation, Autologous