Cellular Environments Alter Performance of rhBMP-2 and Induce Pseudoarthosis**

Background Context: It is still uncertain what factors may intensify or mitigate the osteoinductivity of Bone Morphogenetic Proteins (BMPs) in the clinical situation. As the indications and use of BMPs expand in spinal surgery, it is important to understand how different cellular environments may modulate its effectiveness.

Purpose: This study evaluates the osteoinductivity of rhBMP-2 in different in vivo cellular environments by introducing fibroblast, nucleus pulposus, annulus fibrous, muscle, or marrow cells into the implant site of rats undergoing posterolateral fusion with rhBMP-2.

Patient Sample: 68 female Lewis rats

Methods: Harvest & Culturing Procedure: Adherent skin fibroblasts, annulus fibrosus, nucleus pulposus, muscle, and bone marrow cells were separately harvested from Lewis rats, cultured and expanded and maintained in DMEM. Dosing: Several concentrations within the range of effective doses (ED) of rhBMP2 (0.16, 0.032, 0.006 mg/ml) for osteoinductivity in a rat were screened. The ED100 (0.032 mg/ ml) rhBMP2 at which 100% of the rats were fused was selected for evaluation with various cell types. Surgical Procedure & Treatments: In 60 Lewis rats a posterolateral intertransverse process L4-L5 fusion was performed using one of two doses (0.16mg/ml, 0.032mg/ml) of rhBMP-2 in absorbable collagen sponge (ACS (1.0 x 0.5 x 0.5)/side) combined with one of three quantities (5 x10^6, 10 x10^6 20 x10^6) of the above cell types (n=6 rats each). Control rats (n=4 each) were implanted with the same concentrations of rhBMP-2 with ACS carrier alone and with added media only — no cells (DMEM 0.125ml/side). Rats were sacrificed at 8 weeks. Sites were evaluated using manual testing, radiographs and histology.

Results: S: All rats without implanted cells were completely fused at 0.16 mg rhBMP-2 (4/4), 0.032 mg rhBMP-2/ACS (7/7), and 0.032 mg rhBMP-2/ACS with added media DMEM (4/4). Half the rats were fused at a concentration of 0.006 mg/ml rhBMP-2 (1/2). Fusion was nearly completely inhibited when implanted with any of the quantities of AF, NP, muscle, and marrow cells mixed with a concentration of 0.032 mg rhBMP-2. Fibrous tissue was observed within the osseous mass histologically when not fused. At the higher concentration 0.16 mg rhBMP-2 inhibition was observed with fibroblasts at the 10 x10^6 cell quantity (2/4) and to a lesser degree at 5 x10^6 cell quantity (4/6).

Conclusions: L4-L5 posterolateral pseudarthrosis was experimentally induced by altering the cellular environment in rats at concentrations of rhBMP-2 that consistently induce posterolateral fusion in rats. This study demonstrates that rhBMP-2´s ability to induce a spinal fusion varied as a function of cellular environment. Fibroblasts, disc cells (NP & AF), and muscle cells almost completely inhibited fusion at the lower concentrations of rhBMP-2. At the higher dose of 0.16 mg of rhBMP-2/ACS, fusion was inhibited in a concentration dependent manner with fibroblasts. Marrow cells did not demonstrate any beneficial effect with BMP. This model may allow us to understand inconsistencies and optimize BMP´s performance in human posterolateral spinal fusions.

**The FDA has not cleared a drug and/or medical device the use described in this presentation (i.e., the drug or medical device is being discussed in an “off-label" use).