Nitric Oxide Modulates RHBMP-2 Induced Corticocancellous Autograft Incorporation in Rat Intertransverse Fusion

INTRODUCTION: Nitric oxide (NO) has important physiological functions including the modulation of fracture healing. Recombinant human BMP-2 (rhBMP-2) enhances spinal fusion. With progression of fusion there is a remodelling of the fusion mass bone accompanied with a decrease in the fusion mass size. It is not known whether nitric oxide has a role in spinal fusion or rhBMP-2 enhanced spinal fusion and remodelling.

MATERIALS AND METHODS: We studied this in a novel rat intertransverse fusion model using a defined volume of bone graft (7 caudal vertebrae) along with 157 mm3 of absorbable Type-1 collagen sponge (Helistat ®) carrier, which was compacted and delivered using a custom jig for achieving a similar graft density from sample to sample. The control groups consisted of a sham operated group (S, n=20), an autograft + carrier group (AC, n=28) and a group consisting of 43µg of rhBMP-2 (Genetics Institute, Andover, MA) mixed with autograft + carrier (ACB, n=28). Two experimental groups received a nitric oxide synthase (NOS) inhibitor, NG-nitro L-arginine methyl ester (L-NAME, Sigma Chemicals, St Louis, MO) in a dose of 1mg/ml ad lib in the drinking water (ACL, n=28) and one of these experimental groups had rhBMP-2 added to the graft mixture at the time of surgery (ACLB, n=28). Rats were sacrificed at 22 days and 44 days, spinal columns dissected and subjected to high density radiology (faxitron), biomechanics and undecalcified histology.

RESULTS: On a radiographic score (0-4) indicating progressive maturation of bone fusion mass, no difference was found between the AC and ACL groups, however, there was a significant enhancement of fusion when rhBMP-2 was added (ACB group,3.3±0.2) when compared to the AC group (1±0) (p<.001). However, on day 44, the ACLB group (3.3±0.2) showed significantly less fusion progression when compared to the ACB group (4±0) (p<0.01). There was a 25% (p<0.05) more fusion-mass-area in day 44 of ACLB group (297±26 mm3) when compared to day 44 of the ACB group (225±16 mm3) indicating that NOS inhibition delayed the remodelling of the fusion mass. Biomechanically, the rhBMP-2 groups were stiffer at all time points compared to the NOS inhibited groups. Undecalcified histology demonstrated that there was a delay in graft incorporation whenever NOS was inhibited (ACL and ACLB groups) as assesed by a 5 point histological maturation score.

CONCLUSIONS: We have demonstrated in a novel model of rat intertransverse process fusion that nitric oxide synthase modulates rhBMP-2 induced corticocancellous autograft incorporation.