Fast degradable citrate-based bone scaffold promotes spinal fusion

Abstract

It is well known that high rates of fusion failure and pseudoarthrosis development (5~35%) are concomitant in spinal fusion surgery, which was ascribed to the shortage of suitable materials for bone regeneration. Citrate was recently recognized to play an indispensable role in enhancing osteconductivity and osteoinductivity, and promoting bone formation. To address the material challenges in spinal fusion surgery, we have synthesized mechanically robust and fast degrading citrate-based polymers by incorporating N-methyldiethanolamine (MDEA) into clickable poly(1, 8-octanediol citrates) (POC-click), referred to as POC-M-click. The obtained POC-M-click were fabricated into POC-M-click-HA matchstick scaffolds by compositing with hydroxyapatite (HA) for interbody spinal fusion in a rabbit model. Spinal fusion was analyzed by radiography, manual palpation, biomechanical testing, and histological evaluation. At 4 and 8 weeks post surgery, POC-M-click-HA scaffolds presented optimal degradation rates that facilitated faster new bone formation and higher spinal fusion rates (11.2±3.7, 80±4.5 at week 4 and 8, respectively) than the poly(L-lactic acid)-HA (PLLA-HA) control group (9.3±2.4 and 71.1±4.4) (p<0.05). The POC-M-click-HA scaffold-fused vertebrates possessed a maximum load and stiffness of 880.8±14.5 N and 843.2±22.4 N/mm, respectively, which were also much higher than those of the PLLA-HA group (maximum: 712.0±37.5 N, stiffness: 622.5±28.4 N/mm, p<0.05). Overall, the results suggest that POC-M-click-HA scaffolds could potentially serve as promising bone grafts for spinal fusion applications.

Figure 1

(A) Synthesis of N-methyldiethanolamine (MDEA) modified poly(1, 8-octanediol citrate)-click (POC-M-click) pre-polymers: pre-POC-M-N3 and pre-POC-Al. (B) Fabrication process of POC-M-click-HA matchstick bone scaffold.

Figure 2

Implantation of scaffolds in a rabbit interbody fusion model. (A) POC-M-click-HA and PLLA-HA scaffolds prior to implantation. Scaffold size: 2 mm × 2 mm × 10 mm. (B and C) Pictures taken during the operation. (D) Visual observation of successful fusion at 12 weeks. Arrows point to fused vertebrae.

Figure 3

(A) Degradation profiles of POC-M-click polymer films in PBS (pH 7.4) compared with POC and POC-click polymer films. All polymers were crosslinked at 100°C for 3 days. Representative SEM images of POC-M-click-HA porous matchstick scaffold fabricated with (B) and without (C) using PVP for salt bonding.

Figure 4

Scanning electron microscopy (SEM) images of bone marrow-derived mesenchymal stem cells (BMSCs) grown on the POC-M-click-HA scaffolds (3 days) taken at different magnifications: (A) 200× and (B) 1000×.

Figure 5

Radiography observation of spinal fusion. (A and B) Representative X-ray images of specimens at 4 and 8 weeks after surgery. Black arrows indicate the fusion segment. (C) Fusion rate at 4, 8, and 12 weeks (**p<0.05, *p>0.05). (D) Representative micro-CT scan along the sagittal plane and (E) Three-dimensional reconstruction of a specimen after 12 weeks of surgery with new bone indicated by red arrows.

Figure 6

The maximum bending load (A) and stiffness (B) of the fusion segment 12 weeks after surgery. (**p<0.05).

Figure 7

Photomicrographs of H&E-stained (A and C) and Masson’s trichrome-stained (B and D) tissue sections of the explanted PLLA-HA (A and B) and POC-M-click-HA scaffolds (C and D) 4, 8, and 12 weeks after operation. Yellow arrows indicate vacuoles formed upon material degradation and black arrows indicate the residual unabsorbed materials. Green arrows indicate a large amount of new bone formation surrounding the materials.

Figure 8

Photomicrographs of Safranin O/Fast green stained tissue sections of the explanted PLLA-HA (A and B) and POC-M-click-HA scaffolds (C and D) at 4 and 8 weeks after operation. Yellow arrows indicate the cartilage formed surrounding the materials and red arrows indicate the residual unabsorbed materials.