Retroviral delivery of Noggin inhibits the formation of heterotopic ossification induced by BMP-4, demineralized bone matrix, and trauma in an animal model.
BACKGROUND
The heterotopic ossification of muscles, tendons, and ligaments is a common problem faced by orthopaedic surgeons. We investigated the ability of Noggin (a BMP [bone morphogenetic protein] antagonist) to inhibit heterotopic ossification.


METHODS
Part 1: A retroviral vector carrying the gene encoding human Noggin was developed and used to transduce muscle-derived stem cells. Part 2: Cells transduced with BMP-4 were implanted into both hind limbs of mice along with either an equal number, twice the number, or three times the number of Noggin-expressing muscle-derived stem cells (treated limb) or with nontransduced muscle-derived stem cells (control limb). At four weeks, the mice were killed and radiographs were made to look for evidence of heterotopic ossification. Part 3: Eighty milligrams of human demineralized bone matrix was implanted into the hind limbs of SCID (severe combined immunodeficiency strain) mice along with 100,000, 500,000, or 1,000,000 Noggin-expressing muscle-derived stem cells (treated limbs) or nontransduced muscle-derived stem cells (control limbs). At eight weeks, the mice were killed and radiographs were made. Part 4: Immunocompetent mice underwent bilateral Achilles tenotomy along with the implantation of 1,000,000 Noggin-expressing muscle-derived stem cells (treated limbs) or nontransduced muscle-derived stem cells (control limbs). At ten weeks, the mice were killed and radiographs were made.


RESULTS
Part 1: An in vitro BMP inhibition assay demonstrated that Noggin was expressed by muscle-derived stem cells at a level of 280 ng per million cells per twenty-four hours. Part 2: Three varying doses of Noggin-expressing muscle-derived stem cells inhibited the heterotopic ossification elicited by BMP-4-expressing muscle-derived stem cells. Heterotopic ossification was reduced in a dose-dependent manner by 53%, 74%, and 99%, respectively (p < 0.05). Part 3: Each of three varying doses of Noggin-expressing muscle-derived stem cells significantly inhibited the heterotopic ossification elicited by demineralized bone matrix. Heterotopic ossification was reduced by 91%, 99%, and 99%, respectively (p < 0.05). Part 4: All eleven animals that underwent Achilles tenotomy developed heterotopic ossification at the site of the injury in the control limbs. In contrast, the limbs treated with the Noggin-expressing muscle-derived stem cells had a reduction in the formation of heterotopic ossification of 83% and eight of the eleven animals had no radiographic evidence of heterotopic ossification (p < 0.05).


CONCLUSIONS
The delivery of Noggin mediated by muscle-derived stem cells can inhibit heterotopic ossification caused by BMP-4, demineralized bone matrix, and trauma in an animal model.


CONCLUSIONS
Gene therapy to deliver Noggin may become a powerful method to inhibit heterotopic ossification in targeted areas of the body.
Hannallah
David
D
Department of Orthopaedic Surgery, University of Pittsburgh, PA 15213-2582, USA.
Peng
Hairong
H
Young
Brett
B
Usas
Arvydas
A
Gearhart
Brian
B
Huard
Johnny
J
D000125:Achilles Tendon / Q000601:surgery; D000256:Adenoviridae; D000818:Animals; D001857:Bone Matrix / Q000637:transplantation*; D055415:Bone Morphogenetic Protein 4; D019485:Bone Morphogenetic Proteins / Q000378:metabolism / Q000494:pharmacology*; D002352:Carrier Proteins; D004195:Disease Models, Animal; D018014:Gene Transfer Techniques; D016014:Linear Models; D051379:Mice; D008810:Mice, Inbred C57BL; D016513:Mice, SCID; D018482:Muscle, Skeletal / Q000166:cytology / Q000378:metabolism / Q000637:transplantation; D009999:Ossification, Heterotopic / Q000517:prevention & control*; D011506:Proteins / Q000378:metabolism / Q000494:pharmacology*; D000099176:Noggin Protein
D055415:Bone Morphogenetic Protein 4; D019485:Bone Morphogenetic Proteins; D002352:Carrier Proteins; D011506:Proteins; D000099176:Noggin Protein; C525809:BMP4 protein, human; C525810:Bmp4 protein, mouse