Tissue-Engineered Bone With 3-Dimensionally Printed beta-Tricalcium Phosphate and Polycaprolactone Scaffolds and Early Implantation: An In Vivo Pilot Study in a Porcine Mandible Model

Abstract

Purpose: Deep bone penetration into implanted scaffolds remains a challenge in tissue engineering. The purpose of this study was to evaluate bone penetration depth within 3-dimensionally (3D) printed beta-tricalcium phosphate (beta-TCP) and polycaprolactone (PCL) scaffolds, seeded with porcine bone marrow progenitor cells (pBMPCs), and implanted early in vivo.

Materials and Methods: Scaffolds were 3D printed with 50% beta-TCP and 50% PCL. The pBMPCs were harvested, isolated, expanded, and differentiated into osteoblasts. Cells were seeded into the scaffolds and constructs were incubated in a rotational oxygen-permeable bioreactor system for 14 days. Six 2- x 2-cm defects were created in eachmandible (N= 2 minipigs). In total, 6 constructs were placed within defects and 6 defects were used as controls (unseeded scaffolds, n = 3; empty defects, n = 3). Eight weeks after surgery, specimens were harvested and analyzed by hematoxylin and eosin (H&E), 4',6-diamidino-2-phenylindole (DAPI), and CD31 staining. Analysis included cell counts, bone penetration, and angiogenesis at the center of the specimens.

Results: All specimens (N = 12) showed bone formation similar to native bone at the periphery. Of 6 constructs, 4 exhibited bone formation in the center. Histomorphometric analysis of the H& E-stained sections showed an average of 22.1% of bone in the center of the constructs group compared with 1.87% in the unseeded scaffolds (P <.05). The 2 remaining constructs, which did not display areas of mature bone in the center, showed massive cell penetration depth by DAPI staining, with an average of 2,109 cells/0.57 mm(2) in the center compared with 1,114 cells/0.57 mm(2) in the controls (P <.05). CD31 expression was greater in the center of the constructs compared with the unseeded scaffolds (P <.05).

Conclusion: 3D printed beta-TCP and PCL scaffolds seeded with pBMPCs and implanted early into porcine mandibular defects display good bone penetration depth. Further study with a larger sample and larger bone defects should be performed before human applications.