[Repair of alveolar bone defect with tissue engineered bone: an experimental study of dogs].
OBJECTIVE
To study the feasibility of repairing experimental horizontal alveolar bone defects by tissue engineering based on bone marrow stromal cells (BMSC).


METHODS
Dog bone marrow mononuclear cells were isolated from the bone marrow by gradient centrifugation and then cultured in conditional medium to be induced to become osteogenic. Immunohistochemistry was used to examine the expression of core-binding factor alpha subunit 1 (Cbfa1), osteocalcin (OCN), and type I collagen in the cultured BMSCs. Histochemical technique was used to examine the expression of alkaline phosphatase (AKP) in the BMSCs. Inversed phase-contrast microscopy and electron microscopy were used to observe the morphology and proliferation of the BMSCs. Induced BMSCs at passage 3 were harvested and mixed with calcium alginate to form a gelatin form cell-scaffold construct. A horizontal alveolar bone defect (5 mm high) was created surgically in each buccal side of the mandibular premolars 3 and 4 and molar 1 of 11 dogs. The defects was randomly repaired with a cell-scaffold construct (experimental group, 20 teeth), calcium alginate alone (control group A, 15 teeth), or left untreated (control group B, 12 teeth). At four, twelve, and twenty-four weeks after operation, 2, 7, 2 dogs were killed respectively and block sections of mandibular bones at the defects were collected and processed for gross and histological observation as well as X-ray examination. The status of bone repair 12 weeks after operation in the 3 groups was compared.


RESULTS
In vitro induced BMSCs exhibited an osteogenic phenotype. Since the passage 3 calcium salt sedimentation could be seen in the extracellular stroma of BMSCs. Cbfa1, type I collagen, and AKP were expressed in the BMSCs in every passage. OCN was expressed since the second passage. Histologically, bone nodule structure was observed in the experimental group 4 weeks after operation. The engineered bone became more mature, similar to the normal bone, 12 weeks after operation. Twelve weeks after operation, the alveolar ridge regeneration amounted to a repair height of 2.43 +/- 0.93 mm, 0.98 +/- 0.87 mm, and 0.78 +/- 0.75 mm and reached 48.59%, 19.74%, and 15.76% of the original height in the experimental group, control group A, and control group B respectively, with a significant difference between the experimental and control groups A and B (all P < 0.01).


CONCLUSIONS
BMSCs can be induced to become osteogenic and be used as seed cells to engineer bone tissue and repair experimental alveolar bone defect.
Wang
Min
M
Department of Plastic Surgery of the 9th Hospital, Shanghai Second Medical University, Shanghai Tissue Engineering Research Center, Shanghai 200011, China.
Weng
Yu-lai
YL
Hu
Xiao-jie
XJ
Zhang
Yan
Y
Chai
Gang
G
Zhu
Lian
L
Liu
Wei
W
Cui
Lei
L
Feng
Xi-ping
XP
Cao
Yi-lin
YL
D000469:Alkaline Phosphatase / Q000032:analysis; D016301:Alveolar Bone Loss / Q000628:therapy*; D000818:Animals; D001854:Bone Marrow Cells / Q000166:cytology*; D002454:Cell Differentiation; D024042:Collagen Type I / Q000032:analysis; D050659:Core Binding Factor Alpha 1 Subunit; D050660:Core Binding Factors; D004285:Dogs; D005260:Female; D008297:Male; D009363:Neoplasm Proteins; D015675:Osteocalcin / Q000032:analysis; D010012:Osteogenesis; D017154:Stromal Cells / Q000502:physiology; D023822:Tissue Engineering; D014157:Transcription Factors / Q000032:analysis
D024042:Collagen Type I; D050659:Core Binding Factor Alpha 1 Subunit; D050660:Core Binding Factors; D009363:Neoplasm Proteins; D014157:Transcription Factors; D015675:Osteocalcin; D000469:Alkaline Phosphatase