An animal model study for bone repair with encapsulated differentiated osteoblasts from adipose-derived stem cells in alginate

Document Type : Original Article

Authors

1 Department of Anatomical Sciences, Medical School, Isfahan University of Medical Sciences, Isfahan, Iran

2 Cancer Prevention Research Center, Medical School, Isfahan University of Medical Sciences, Isfahan, Iran Institute of Novin Tahlilgarane Nesfe-jahan, Isfahan, Iran

Abstract

Objective(s): Adipose derived stem cells (ADSCs) can be engineered to express bone specific markers. The aim of this study is to evaluate repairing tibia in animal model with differentiated osteoblasts from autologous ADSCs in alginate scaffold.
Materials and Methods: In this study, 6 canine’s ADSCs were encapsulated in alginate and differentiated into osteoblasts. Alkaline phosphatase assay (ALP) and RT-PCR method were applied to confirm the osteogenic induction. Then, encapsulated differentiated cells (group 1) and cell-free alginate (group 2) implanted in defected part of dog's tibia for 4 and 8 weeks. Regenerated tissues and compressive strength of samples were evaluated by histological and Immunohistochemical (IHC) methods and Tensometer Universal Machine.
Results: Our results showed that ADSCs were differentiated into osteoblasts in vitro, and type I collagen and osteocalcin genes expression in differentiated osteoblasts was proved by RT-PCR. In group 2, ossification and thickness of trabecula were low compared to group 1, and in both groups woven bone was observed instead of control group's compact bone. Considering time, we found bone trabeculae regression and ossification reduction after 8 weeks compared with 4 weeks in group 2, but in group 1 bone formation was increased in 8 weeks. Presence of differentiated cells caused significantly more compressive strength in comparison with group 2 (P-value ≤0.05).
Conclusion: This research showed that engineering bone from differentiated adipose-derived stem cells, encapsulated in alginate can repair tibia defects.

Keywords


1. Mankin HJ, Hornicek FJ, Raskin KA. Infection in massive bone allograft. Clin Orthop Relat Res 2005; 432:210–216.
2. Nishida J, Shimamura T. Methods of reconstruction for bone defect after tumor excision: a review of alternatives. Med Sci Monit 2008; 14:107–113.
3. Hou CH, Yang RS, Hou SM. Hospital-based allogenic bone bank 10 year experience. J Hosp Infect 2005; 59:41–45.
4. Dormer NH, Busaidy K, Berkland CJ,  Detamore  MS. Osteochondral interface regeneration of rabbit mandibular condyle with bioactive signal gradients. J Oral Maxillofac Surg 2011; 69:e50-57
5. Mastrogiacomo M, Muraglia A, Komlev V,Peyrin F, Rustichelli F. Tissue engineering of bone: search for a better scaffold. Orthod Craniofac Res 2005; 8:277-284.
6. Jin QM, Takita H, Kohgo T, Atsumi K, Itoh H, Kuboki Y. Effects of geometry of hydroxyapatite as a cell substratum in BMP-induced ectopic bone formation. J Biomed Mater Res 2000; 52:491-499.
7. Boden SD. Bioactive factors for bone tissue engineering. Clin Orthop Relat Res 1999; 367:84-94.
8. Whang K, Goldstick TK, Healy KE. A biodegradable polymer scaffold for delivery of osteotropic factors. Biomaterials 2000; 21:2545-2551.
9.  Lee SC, Shea M, Battle MA, Kozitza K, Ron E, Turek T, et al. Healing of large segmental defects in rat femurs is aided by RhBMP-2 in PLGA matrix. J Biomed Mater 1994; 28:1149-1156.
10. Cai X , LinY, Ou G, Luo E, ManY, Yuan Q, et al. Ectopic osteogenesis and chondrogenesis of bone marrow stromal stem cells in alginate system. Cell Biol Int  2007; 31:776-783.
11. Williams CG, Kim TK, Taboas A, Malik A, Manson P, Elisseeff J. In vitro chondrogenesis of bone marrow-derived mesenchymal stem cells in a photopolymerizing hydrogel. Tissue Eng 2003; 9:679-688.
12. Murtas S, Capuani G, Dentini M, Manetti C, Masci G, Massimi M, et al. Alginate beads as immobilization matrix for hepatocytes perfused in a bioreactor: a physico-chemical characterization. J Biomater Sci Polym Ed 2005; 16:829-846.
14. Abbah SA, Lu EW, Chan ED, Cheung EK. Osteogenic behavior of alginate encapsulated bone marrow stromal cells: an in vitro study. J Mater Sci Mater Med 2008; 19:2113-2119.
15. Liao W, Yang Z, Deng L, Li X, Sun T, Luo J. Morphological and biomechanical study on in vivo osteogenesis after repair of cranial defects with plastic engineered bone in rabbits. Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi 2005; 19:460-463.
16. Alsberg H, Yang z. Regulating Bone Formation via Controlled Scaffold Degradation. J Bone Miner Res 2003; 18:923-929.
17.  Zuk PA, Zhu M, Ashjian P, De Ugarte DA, Huang JI, Mizuno H, et al. Human adipose tissue is a source of multipotent stem cells. Molecul Biol 2002; 13:4279-4295.
18. Gomillion CT, Burg KJ. Stem cells and adipose tissue engineering. Biomaterials 2006; 27:6052-6063.
19. Halvorsen YD, Franklin D, Bond AL, Hitt DC, Auchter C, Boskey AL, et al. Extracellular matrix mineralization and osteoblast gene expression by human adipose tissue-derived stromal cells. Tissue Eng  2001; 7:729-741.
20. Hashemibeni B, Goharian V, Esfandiari E, Sadeghi F, Fasihi F, Alipur R, et al. An animal model study for repair of tracheal defects with autologous stem cells and differentiated chondrocytes from adipose-derived stem cells. J Pediatr Surg 2012; 47:997-2003.
21. Birmingham E, Niebur GL, McHugh PE, Shaw G, Barry FP, McNamara LM.  Osteogenic differentiation of mesenchymal stem cells is regulated by osteocyte and osteoblast cells in a simplified bone nich. Europ Cells Mat 2012; 23:13-27.
22. Hashemibeni B, Razavi Sh, Esfandiary E, Karbasi S, Mardani M, Nasresfahani M. Induction of chondrogenic differentiation of human adipose-derived stem cells with TGF-b3 pellet culture system. Iran J Basic Med Sci 2008; 11:10-17.
23. Hardingham T. Tissue engineering: Chondrocytes and cartilage. Arthritis Res 2002; 4:63-68.
24. Diduch DR, Jordan LC, Mierisch CM, Balian G. Marrow stromal cells embedded in alginate for repair of osteochondral defects. Arthroscopy 2000; 16:571-577.
25. Mardani M, Hashemibeni B, Ansar M, Zarkesh SH, Kazemi M, GoharianV, et al. Comparison between chondrogenic markers of differentiated chondrocytes from adipose derived stem cells and articular chondrocytes in vitro. Iran J Basic Med Sci 2013; 16:764-771.
26. Zhensheng Li, Hassna R, Kip D. Chitosan-alginate hybrid scaffoldfor bone tissue engineering. Biomaterial 2005; 26:3919-3928.
27.  Majmudar G, Bole D, Goldstein SA. Bone cell culture in a three dimensional polymer bead stabilizes the differentiated phenotype and evidence that osteoblastic cells synthesize type III collagen and
fibronectin. J Bone Miner Res 1991; 6:837-839.
28. LinY, Tang W, Ling Wu, Jing W, Li X, Wu Y, et al. Bone regeneration by BMP-2 enhanced adipose stem cells loading on alginate gel. Histochem Cell Biol 2008; 129:203-210.
29. Li Z, Ramay HR, Hauch KD, Xiao D, Zhang M. Chitosan-alginate hybrid scaffolds for bone tissue engineering. Biomaterials  2005; 26:3919-3928.