Posterolateral Spinal Fusion in Rabbits Using a RP-based PLGA/ TCP/Col/BMSCs-OB Biomimetic Grafting Material

Abstract

Three-dimensional highly porous poly(DL-lactic-co-glycolic acid)/ tricalcium phosphate (PLGA/TCP) scaffolds were fabricated using a rapid prototyping technique (RP). The 3D rhombic lamellar PLGA/TCP carriers (20 mm × 20 mm × 3 mm) subsequently were coated with collagen type I (Col) to produce PLGA/TCP/Col composites. Both the RP-based PLGA/TCP scaffolds and the PLGA/TCP/Col composites were observed by scanning electron microscopy. Forty New Zealand white rabbits were equally randomized into 2 groups (group A and group B) and bilaterally underwent posterolateral intertransverse process arthrodesis at the L4—L5 level using the following graft materials: In group A, PLGA/TCP/Col/BMSCs-OB composites (on the right side, group A1, n = 20) and autogenous iliac bone grafts (on the left side, group A2, n = 20) were used; In group B, PLGA/TCP scaffolds plus fresh autogenous bone marrow (on the right side, group B1, n = 20) and PLGA/TCP scaffolds alone (on the left side, group B2, n = 20) were utilized. In group A1, rabbit bone marrow stromal cells (BMSCs) were isolated and cultured under the osteogenic conditions (BMSCs-OB). Structural PLGA/TCP/Col composites then were efficiently loaded with BMSCs-OB and cultured 5 days to make PLGA/TCP/ Col/BMSCs-OB biomaterials. Rabbits were sacrificed after 12-week follow-up and the spinal fusion were evaluated by a general observation, a manual palpation test, histological analyses and radiography. As a result, RP established PLGA/TCP scaffolds with appropriate biomaterial properties including satisfactory microstructure, inter-connectivity and porosity. Modifications to the structural highly porous PLGA/TCP scaffolds with Col (PLGA/TCP/Col) essentially increased the affinity of the carriers to seeding cells. In group A1, radiological evaluation revealed strong ability of new bone formation and bony fusion in the implanted sites and histological analyses showed highly cellular bone marrow between the newly formed trabecular bone was present in the fusion mass. In group A2, there was a reduced amount of newly formed bone. In group B1, only a few bony fusions were obtained. In group B2, PLGA/TCP scaffolds were biocompatible and biodegradable; whereas, no newly formed bone or bony fusion was found. Twelve weeks after surgery, spinal fusion rates in groups of A1, A2, B1, and B2 were 70.0%(14/20), 45.0%(9/20), 15.8%(3/19), and 0%(0/19), respectively. The rates of fusion were significantly higher in groups of A1 and A2 compared with groups of B1 and B2 (p<0.01), and there was no significant difference of fusion rate between group A1 and group A2 (p>0.05). Therefore, RP-based 3D PLGA/TCP/Col/BMSCs-OB biomaterial holds promise as a bone grafting substitute for spinal fusion. Our attempts may provide a novel method for biofabrication of the bionic construct.

Keywords

rapid prototyping (RP)scaffold bone marrow stromal cells (BMSCs)spine fusion rabbit tissue engineering.

Get full access to this article

View all access options for this article.

References

Kyriakidou, K. , Lucarini, G. , Zizzi , A. , Salvolini, E. , Belmonte, M.M. , Mollica, F. et al. (2008 ). Dynamic Co-seeding of Osteoblast and Endothelial Cells on 3D Polycaprolactone Scaffolds for Enhanced Bone Tissue Engineering, J. Bioact. Comp. Polym., 23(3): 227-243.

Lopez-Heredia, M.A. , Sohier, J. , Gaillard , C. , Quillard, S. , Dorget, M. and Layrolle, P. (2008). Rapid Prototyped Porous Titanium Coated with Calcium Phosphate as a Scaffold for Bone Tissue Engineering , Biomaterials, 29(17): 2608-2615.

Xiong, Z. , Yan, Y. , Wang, S. , Zhang, R. and Zhang, C. ( 2002). Fabrication of Porous Scaffolds for Bone Tissue Engineering via Low-temperature Deposition, Scripta. Mater. , 46(11): 771-776.

Liu, X. and Ma, P.X. ( 2004). Polymeric Scaffolds for Bone Tissue Engineering , Ann. Biomed. Eng., 32(3): 477-486.

Robey, P.G. , Kuznetsov, S.A. , Riminucci, M. and Bianco, P. ( 2007). Skeletal (‘‘Mesenchymal’’) Stem Cells for Tissue Engineering, Methods. Mol. Med., 140: 83-99.

Derubeis, A.R. and Cancedda, R. (2004). Bone Marrow Stromal Cells (BMSCs) in Bone Engineering: Limitations and Recent Advances , Ann. Biomed. Eng., 32(1): 160-165.

Ma, X. , Hu, Y. , Yan, Y. , Xiong, Z. , Lü, R. , Wang, J. et al. (2006). The Experimental Study of Biomimetic Artificial Cartilage Fabrication In Vitro and Ectopic Chondrogenesis In Vivo, Sheng. Wu. Yi. Xue. Gong. Cheng. Xue. Za. Zhi., 23(4): 795-799.

Ma, X. , Hu, Y.Y. , Wu, X.M. , Liu, J. , Xiong, Z. , Yan, Y.N. et al. (2009). Fabrication of a RP-based Biomimetic Grafting Material for Bone Tissue Engineering, Mater. Sci. Forum., 626-627: 553-558.

Machado, C.B. , Ventura, J.M. , Lemos, A.F. , Ferreira, J.M. , Leite, M.F. and Goes, A.M. (2007). 3D Chitosan-gelatin-chondroitin Porous Scaffold Improves Osteogenic Differentiation of Mesenchymal Stem Cells, Biomed. Mater. , 2(2): 124-131.

10.

Xu, W. , Wang, X. , Yan, Y. and Zhang, R. ( 2008). Rapid Prototyping of Polyurethane for the Creation of Vascular Systems, J. Bioact. Comp. Polym., 23(2): 103-114.

11.

Ma, X. , Wu, X. , Hu, Y. , Xiong, Z. , Lv, R. , Wang, J. et al. (2009). Intervertebral Spinal Fusion Using a RP-based PLGA/TCP/bBMP Biomimetic Grafting Material , J. Bioact. Comp. Polym., 24(1 Suppl): 146-157.

12.

Liu, G. , Hu, Y.Y. , Zhao, J.N. , Wu , S.J. , Xiong, Z. and Lu , R. ( 2004). Effect of Type I Collagen on the Adhesion, Proliferation, and Osteoblastic Gene Expression of Bone Marrow-derived Mesenchymal Stem Cells, Chin. J. Traumatol., 7(6): 358-362.

13.

Huang, W. , Carlsen, B. , Wulur, I. , Rudkin, G. , Ishida, K. , Wu, B. et al. (2004). BMP-2 Exerts Differential Effects on Differentiation of Rabbit Bone Marrow Stromal Cells Grown in Two-dimensional and Three-dimensional Systems and is Required for In Vitro Bone Formation in a PLGA Scaffold, Exp. Cell. Res., 299(2): 325-334.

14.

Sasso, R.C. , LeHuec, J.C. and Shaffrey, C. ( 2005). Iliac Crest Bone Graft Donor Site Pain after Anterior Lumbar Interbody Fusion: A Prospective Patient Satisfaction Outcome Assessment , J. Spinal. Disord. Tech., 18(Suppl): S77-81.

15.

Rhee, J.M. , Patel, N. , Yoon, S.T. and Franklin, B. ( 2007). High Graft Resorption Rates with Dense Cancellous Allograft in Anterior Cervical Discectomy and Fusion, Spine , 32(26): 2980-2984.

16.

Glowacki, J. and Mizuno, S. (2008). Collagen Scaffolds for Tissue Engineering, Biopolymers, 89(5): 338-344.

17.

Kanczler, J.M. and Oreffo, R.O. (2008). Osteogenesis and Angiogenesis: the Potential for Engineering Bone, Eur. Cell. Mater., 15: 100-114.

18.

Yan, Y. , Wang, X. , Xiong , Z. , Liu , H. , Liu , F. , Lin , F. et al. (2005). Direct Construction of a Three-dimensional Structure with Cells and Hydrogel , J. Bioact. Comp. Polym., 20(3): 259-269.

19.

Zhang, T. , Yan, Y. , Wang, X. , Xiong, Z. , Lin, F. , Wu, R. et al. (2007). Three-dimensional Gelatin and Gelatin/Hyaluronan Hydrogel Structures for Traumatic Brain Injury, J. Bioact. Comp. Polym., 22(1): 19-29.

20.

Xu, W. , Wang, X. , Yan, Y. , Zheng, W. , Xiong, Z. , Lin, F. et al. (2007). Rapid Prototyping Three-Dimensional Cell/Gelatin/Fibrinogen Constructs for Medical Regeneration, J. Bioact. Comp. Polym. , 22(4): 363-377.

21.

Yao, R. , Zhang, R. , Yan , Y. and Wang, X. (2009). In Vitro Angiogenesis of 3D Tissue Engineered Adipose Tissues, J. Bioact. Comp. Polym., 24(1): 5-24.

22.

Xiong, Z. , Yan, Y. , Zhang, R. and Wang, X. ( 2005). Organism Manufacturing Engineering Based on Rapid Prototyping Principles, Rapid Prototyping J., 11(3): 160-166.

23.

Ma, X. , Hu, Y. , Wu, X. , Yan, Y. , Xiong, Z. , Lu, R. et al. (2008). Surface Modification of Biodegradable Polymer/TCP Scaffolds and Related Research, Sheng. Wu. Yi. Xue. Gong. Cheng. Xue. Za. Zhi., 25(3): 571-577.

24.

Miao, X. , Tan, D.M. , Li, J. , Xiao, Y. and Crawford, R. ( 2008). Mechanical and Biological Properties of Hydroxyapatite/Tricalcium Phosphate Scaffolds Coated with Poly(Lactic-Co-Glycolic Acid), Acta Biomater., 4(3): 638-645.

25.

Kawazoe, N. , Lin, X. , Tateishi , T. and Chen, G. ( 2009). Three-dimensional Cultures of Rat Pancreatic RIN-5F Cells in Porous PLGA-Collagen Hybrid Scaffolds, J. Bioact. Comp. Polym., 24(1): 25-42.

26.

Minamide, A. , Yoshida, M. , Kawakami , M. , Okada, M. , Enyo, Y. , Hashizume, H. et al. (2007 ). The Effects of Bone Morphogenetic Protein and Basic Fibroblast Growth Factor on Cultured Mesenchymal Stem Cells for Spine Fusion, Spine, 32(10): 1067-1071.

27.

Yee, A.J. , Bae, H.W. , Friess, D. , Roth, S.M. , Whyne, C. , Robbin, M. et al. (2006 ). The Use of Simvastatin in Rabbit Posterolateral Lumbar Intertransverse Process Spine Fusion, Spine. J., 6(4): 391-396.

28.

Erickson, B.P. , Pierce, A.R. , Simpson, A.K. , Nash, J. and Grauer, J.N. (2008). 125I-labeled OP-1 Is Locally Retained in a Rabbit Lumbar Fusion Model, Clin . Orthop. Relat. Res., 466(1): 210-215.