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Tissue-engineered Bone Repair of Sheep Femur Defects with Induced Bone Marrow Stromal Cells
| Content Provider | Semantic Scholar |
|---|---|
| Copyright Year | 2002 |
| Abstract | Introduction: Large bone defect remains difficult to treat because of limited availability of autologous bone grafts. In contrast, tissueengineering technique can generate large bone tissue using a small amount of autologous cells and therefore, provide an optimal solution. Bone marrow stromal cells (BMSCs) have the potential for multilineage (including osteogenic) differentiation and thus are the ideal seed cells for bone tissue engineering. We have successfully engineered flat bone tissue to repair cranial defect using autologous BMSCs in a previously study. In addition, BMSCs can be induced to become osteogenic and produced a high level of ALP, osteocalcin and collagen I. The objective of this study was to investigate the feasibility of engineering long bone tissue and repairing a critical sized femoral bone defect in a sheep model. Methods: Bone marrows were harvested from 12 adult sheep respectively and BMSCs were isolated by their adhesion to culture dish. Cells were in vitro expanded and induced for osteogenic differentiation with 10M dexamethasone and 10mM ß-phospherglycerol added in DMEM plus 10%FBS. The coral (Hainan, China) with a porosity of 4050% and pore sizes ranging from 150 to 200μm was used as the biomaterial. The induced cells (5-6×10) in third passage were seeded onto a coral construct with a cylinder shape of 25mm in length, 16 mm in out-diameter and 10 mm in inner-diameter. The cell-coral construct was co-cultured for 1 week before in vivo implantation. Cell attachment to and matrix production on the coral construct were evaluated with scanning electromicroscope. For surgical procedure, a bone defect of 25mm-length was created at the mid-portion of right femur in each animal followed by an internal fixation with intramedullary bar and inter-locking nails. The defects were either repaired with cell-coral construct in experimental group (n=6) or with coral construct alone in control group (n=6). Bone healing was monitored by radiograph taken at day 1, and months 1,2,3,4,5,6,7 and 8 post-repair respectively. Radiodensity of defect area in both groups was quantitatively analyzed with image software at the time points of day1 and months 1, 2, 3, 4 and 5 respectively. F-test was used for statistical study and a p-value less than 0.05 was considered as significant different. Three animals of each group were sacrificed at 4 and 8 months post-repair respectively for gross and histological examination. Additionally, in three animals of experimental group, the inter-locking nails were removed at 6 months while maintaining the intramedullary bar for another two months and therefore to provide the repaired femur a mechanical loading to promote bone remodeling. Essential results: Cells adhered to at day 1 and fully spread on coral at day 3. Abundant matrix production and the cell proliferation inside the coral micro-pores were observed at day 7. Radiograph showed that the coral construct was clearly visible at day 1 post-repair in both groups. However, the coral construct was partially degraded at the first month and completely disappeared at the second month in control group. Additionally, limited new bone formed at both ends of the defect was observed from months 3 to 8 post-repair. In experimental group, a relatively lower radiodensity at the defect area was observed at the first and the second months. In addition, callus was formed at the interface between the host bone and the coral cylinder 2 month post-repair. Radiodensity of engineered bone in experimental group continued to increase from months 3 to 6, but remained relatively lower than normal cortex bone. At 8 months, engineered bone reached a radiodensity similar to that of normal femur, which has been remodeled into cortex bone at outer layer by the mechanical loading. There is a statistical significant difference in radiodensity between the two groups (Table 1, p<0.05). Gross observation at 4 months showed that control defect was filled mostly with fibrous tissue although small a small amount of new bone was formed at both ends of the defect. In contrast, the experimental defect was filled with abundant regenerated bone tissue, which was rich in vascularity. At 8 months, the control bone defect was connected with fibrous tissue with an abnormal movement. The experimental defect was completely repaired with tissue-engineered bone, which had white color and firm texture and was able to bear strong stress. More importantly, the repaired femur was fully functional as the animal could walk normally after removing of internal fixation 6 months post-repair. Histology demonstrated that the control defect contained mainly fibrous tissue in which undegraded coral particle remained visible at 4 months, but completely disappeared at 8 months. In contrast, woven or trabecular bone tissue was observed in experimental defect at 4 months, and original or irregular osteons were formed at 8 months post-repair. Discussion: This study demonstrated that long weight-bearing bone tissue can be successfully engineered in sheep using induced BMSCs and coral. Additionally, coral can be fully degraded within two months, which matchs the time for bone regeneration and does not interfere with bone remodeling. Thus, we consider coral a biomaterial better than other slow-degrading material like tricalcium phosphate and hydroxyapatite for bone engineering. Unlike the previously reported study, BMSCs used in this study were in vitro induced to become osteogenic prior to in vivo implantation. In addition, we in vitro cocultured the BMSCs with coral construct for 1 week before transplantation to make sure that BMSCs were well-attached to the coral construct and thus could efficiently proliferate and produce their own matrix after in vivo implantation. These techniques may explain that this study could achieve a much higher success rate (6/6) of bone repair than that (3/7) reported previously.Our next step will focus on mechanical study of engineered long bone tissue. References 1. Shang, Q.X., Wang, Z., Liu, W., Shi, Y.H., Cui, L., and Cao, Y.L. Tissue-engineered bone repair of sheep cranial defects with autologous bone marrow stromal cells. J. Craniofac. Surg. 2001;12:586. 2. Petite H, Viateau V, Bensaid W, Meunier A, de Pollak C, Bourguignon M, Oudina K, Sedel L, Guillemin G. Tissue-engineered bone regeneration. Nat Biotechnol. 2000;18:959. |
| File Format | PDF HTM / HTML |
| Alternate Webpage(s) | https://www.ors.org/Transactions/49/0934.pdf |
| Language | English |
| Access Restriction | Open |
| Content Type | Text |
| Resource Type | Article |
