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. 2022 Jan 1;47(1):82-89.
doi: 10.1097/BRS.0000000000004142.

Preclinical Safety of a 3D-Printed Hydroxyapatite-Demineralized Bone Matrix Scaffold for Spinal Fusion

Mark Plantz  1   2 , Joseph Lyons  1   2 , Jonathan T Yamaguchi  1   2 , Allison C Greene  1   2 , David J Ellenbogen  1   2 , Mitchell J Hallman  1   2 , Vivek Shah  1   2 , Chawon Yun  1   2 , Adam E Jakus  3 , Daniele Procissi  4 , Silvia Minardi  1   2 , Ramille N Shah  2   3 , Wellington K Hsu  1   2 , Erin L Hsu  1   2
Affiliations

Preclinical Safety of a 3D-Printed Hydroxyapatite-Demineralized Bone Matrix Scaffold for Spinal Fusion

Mark Plantz et al. Spine (Phila Pa 1976). .

Abstract

Study design: Prospective, randomized, controlled preclinical study.

Objective: The objective of this study was to compare the host inflammatory response of our previously described hyperelastic, 3D-printed (3DP) hydroxyapatite (HA)-demineralized bone matrix (DBM) composite scaffold to the response elicited with the use of recombinant human bone morphogenetic protein-2 (rhBMP-2) in a preclinical rat posterolateral lumbar fusion model.

Summary of background data: Our group previously found that this 3D-printed HA-DBM composite material shows promise as a bone graft substitute in a preclinical rodent model, but its safety profile had yet to be assessed.

Methods: Sixty female Sprague-Dawley rats underwent bilateral posterolateral intertransverse lumbar spinal fusion using with the following implants: 1) type I absorbable collagen sponge (ACS) alone; 2) 10 μg rhBMP-2/ACS; or 3) the 3DP HA-DBM composite scaffold (n = 20). The host inflammatory response was assessed using magnetic resonance imaging, while the local and circulating cytokine expression levels were evaluated by enzyme-linked immunosorbent assays at subsequent postoperative time points (N = 5/time point).

Results: At both 2 and 5 days postoperatively, treatment with the HA-DBM scaffold produced significantly less soft tissue edema at the fusion bed site relative to rhBMP-2-treated animals as quantified on magnetic resonance imaging. At every postoperative time point evaluated, the level of soft tissue edema in HA-DBM-treated animals was comparable to that of the ACS control group. At 2 days postoperatively, serum concentrations of tumor necrosis factor-α and macrophage chemoattractant protein-1 were significantly elevated in the rhBMP-2 treatment group relative to ACS controls, whereas these cytokines were not elevated in the HA-DBM-treated animals.

Conclusion: The 3D-printed HA-DBM composite induces a significantly reduced host inflammatory response in a preclinical spinal fusion model relative to rhBMP-2.Level of Evidence: N/A.

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Figures

Figure 1.
Figure 1.
Depiction of MRI analysis methodology. Sagittal, axial, and coronal views of an HA-DBM implant are provided both before and after processing. Image processing involved isolating the local edema at the surgical site using threshold-based automatic segmentation of standardized MRI images. The local edema was identified (depicted as red overlay) and the total volume of edema was calculated. The crosshair is at the midline, just superior to the L4/L5 disc. HA-DBM indicates hydroxyapatixte-demineralized bone matrix; MRI, magnetic resonance imaging.
Figure 2.
Figure 2.
Representative axial T2-weighted MR images of animal groups at various time points after surgery are shown (N = 5 per treatment group per time point). A large fluid collection (red arrows) was reproducible in those animals receiving rhBMP-2 that was most pronounced at 2 days after surgical implantation and was not typically seen in the ACS and HA-DBM groups. ACS indicates absorbable collagen sponge; DBM, demineralized bone matrix; HA, hydroxyapatite; MR, magnetic resonance; rhBMP-2, recombinant human bone morphogenetic protein-2.
Figure 3.
Figure 3.
Quantification of inflammatory edema volume within the fusion bed (intramuscular/subfascial) on axial T2-weighted MR images in all animal groups at 2 days, 5 days, 10days, and 8 weeks postoperatively (N = 5 per treatment group per time point). Significantly greater inflammatory edema volume was measured in the rhBMP-2 group as early as 2 days after surgery when compared with the ACS (*) and HA-DBM (^) groups (P < 0.05). ACS indicates absorbable collagen sponge; DBM, demineralized bone matrix; HA, hydroxyapatite; MR, magnetic resonance.
Figure 4.
Figure 4.
Histological evaluation of the implants 2, 5, 10 days and 8 weeks postoperatively, stained with Gill’s hematoxylin, eosin and alcian blue. Yellow arrows in the ACS and rhBMP-2 samples indicate collagen bundles (red, ribbon-like). Yellow arrows in the 2 days HA-DBM sample indicate cells (purple nuclei) in proximity of a DBM particle. Yellow arrows in the 10 days HA-DBM sample indicate residual HA particles. Legend: Scaffold struts = ST; demineralized bone matrix = DBM; new tissue = NT; muscle M; transverse process = TP; trabecular bone = TB. Scale bars: 400 μm. ACS indicates absorbable collagen sponge; DBM, demineralized bone matrix; HA, hydroxyapatite.
Figure 5.
Figure 5.
Serum levels of the following cytokines of interest were quantified longitudinally via ELISA (N = 5 per treatment group per time point): IL-1β, IL-18, TNFα, MIP-1, and MCP-1. Levels were compared among animals treated with ACS alone, rhBMP-2/ACS, and HA-DBM scaffolds. Statistical significance for the HA-DBM group relative to ACS alone (*) and rhBMP-2/ACS (^) groups is indicated (P < 0.05). ACS indicates absorbable collagen sponge; DBM, demineralized bone matrix; ELISA, enzyme-linked immunosorbent assays; HA, hydroxyapatite.
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