Enhanced repair of the anterior cruciate ligament by in situ gene transfer: evaluation in an in vitro model.
The inability of the ruptured anterior cruciate ligament (ACL) of the knee joint to heal spontaneously presents numerous clinical problems. Here we describe a novel, gene-based approach to augment ACL healing. It is based upon the migration of cells from the ruptured ends of the ligament into a collagen hydrogel laden with recombinant adenovirus. Cells entering the gel become transduced by the vector, which provides a basis for the local synthesis of gene products that aid repair. Monolayers of bovine ACL cells were readily transduced by first-generation, recombinant adenovirus, and transgene expression remained high after the cells were incorporated into collagen hydrogels. Using an in vitro model of ligament repair, cells migrated from the cut ends of the ACL into the hydrogel and were readily transduced by recombinant adenovirus contained within it. The results of experiments in which GFP was used as the transgene suggest highly efficient transduction of ACL cells in this manner. Moreover, during a 21-day period GFP+ cells were observed more than 6 mm from the severed ligament. This distance is ample for the projected clinical application of this technology. In response to TGF-beta1 as the transgene, greater numbers of ACL cells accumulated in the hydrogels, where they deposited larger amounts of type III collagen. These data confirm that it is possible to transduce ACL cells efficiently in situ as they migrate from the ruptured ACL, that transduction does not interfere with the cells' ability to migrate distances necessary for successful repair, and that ACL cells will respond in a suitable manner to the products of the transgenes they express. This permits optimism over a possible clinical use for this technology.
Center of Molecular Orthopaedics, Harvard Medical School, Boston, MA 02115, USA.
Arnulf
A
Pascher
Andre F
AF
Steinert
Glyn D
GD
Palmer
Oliver
O
Betz
Jean-Noel
JN
Gouze
Elvire
E
Gouze
Carmencita
C
Pilapil
Stephen C
SC
Ghivizzani
Christopher H
CH
Evans
Martha Meaney
MM
Murray
D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D016118:Anterior Cruciate Ligament / Q000502:physiology; D000070598:Anterior Cruciate Ligament Injuries; D002417:Cattle; D002465:Cell Movement; D002478:Cells, Cultured; D024061:Collagen Type III / Q000032:analysis / Q000378:metabolism; D015870:Gene Expression / Q000235:genetics; D015316:Genetic Therapy / Q000379:methods*; D049452:Green Fluorescent Proteins / Q000032:analysis / Q000235:genetics; D020136:Hydrogel, Polyethylene Glycol Dimethacrylate / Q000494:pharmacology; D012421:Rupture / Q000628:therapy; D014161:Transduction, Genetic / Q000379:methods*; D016212:Transforming Growth Factor beta / Q000032:analysis / Q000235:genetics* / Q000378:metabolism; D053773:Transforming Growth Factor beta1; D014945:Wound Healing
D024061:Collagen Type III; D016212:Transforming Growth Factor beta; D053773:Transforming Growth Factor beta1; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D020136:Hydrogel, Polyethylene Glycol Dimethacrylate