Chinese Journal of Tissue Engineering Research ›› 2021, Vol. 25 ›› Issue (10): 1489-1494.doi: 10.3969/j.issn.2095-4344.3052

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Osteogenesis in vitro and bone defect repair in vivo of bone morphogenetic protein 9 composite collagen based bone repair material

Zhuang Chuanji1, Chen Wenzhao2, Jiang Xinmin1   

  1. 1Department of Orthopedics, Second People’s Hospital of Jingdezhen, Jingdezhen 333000, Jiangxi Province, China; 2Department of Orthopedics, the First Affiliated Hospital of Nanchang University, Nanchang 330006, Jiangxi Province, China
  • Received:2020-05-20 Revised:2020-05-22 Accepted:2020-06-29 Online:2021-04-08 Published:2020-12-17
  • Contact: Jiang Xinmin, Chief physician, Department of Orthopedics, Second People’s Hospital of Jingdezhen, Jingdezhen 333000, Jiangxi Province, China
  • About author:Zhuang Chuanji, Master, Associate chief physician, Department of Orthopedics, Second People’s Hospital of Jingdezhen, Jingdezhen 333000, Jiangxi Province, China
  • Supported by:
    the Project Supported by Jiangxi Health Committee, No. 20204356 (to ZCJ)

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Abstract: BACKGROUND: Collagen-based bone repair materials have been widely used in clinical bone repair due to their excellent osteoconductivity, osteoinduction, degradation, mechanical properties and biocompatibility, but their osteoinduction effect is not as good as that of autogenous bone.
OBJECTIVE: Bone morphogenetic protein 9 (BMP-9) was incorporated into collagen matrix to investigate its osteoinductive ability.
METHODS: The plasmid DNA encoding BMP-9 was loaded into collagen-based bone repair materials to construct a composite of collagen-based bone repair materials. (1) In vitro experiment: The Sprague-Dawley rat adipose mesenchymal stem cells were inoculated with BMP-9 composite collagen-based bone repair material and collagen-based bone repair material. Cell proliferation was detected by MTT assay, and the cell adhesion was observed by scanning electron microscope. Cell adhesion was counted after DAPI staining. Alkaline phosphatase activity was detected by alkaline phosphatase kit. Osteopontin and osteocalcin mRNA expression levels were detected by qRT-PCR. (2) In vivo experiment: Bone defects of 3 mm in diameter and 3 mm in depth were made in the medial epicondyle of the right hind limb of 15 Sprague-Dawley rats. No material was implanted in the blank group. Collagen-based bone repair material was implanted in the control group. BMP-9 composite collagen-based bone repair material was implanted in the observation group. Samples were obtained at 8 weeks after surgery. Micro-CT examination and histological observation were performed. This study was approved by the Ethics Committee of Laboratory Animal Science Center of Nanchang University. 
RESULTS AND CONCLUSION: (1) In vitro experiment showed that there was no significant difference between the proliferation of adipose tissue mesenchymal stem cell cultured for 1, 3, 5, and 7 days in collagen-based bone repair materials compounded with BMP-9 (P > 0.05). (2) After co-culture in vitro for 24 hours, more cells and more pseudopodia appeared on the surface of collagen-based bone repair material compounded with BMP-9. The number and spreading of adherent cells were better than that of collagen-based bone repair material. (3) After 3 and 7 days of osteogenic induction in vitro, the cell alkaline phosphatase activity of the collagen-based bone repair material compounded with BMP-9 was higher than that of the collagen-based bone repair material (P < 0.05). After 7 days of osteogenic induction, osteopontin and osteocalcin mRNA expression was higher than that of collagen-based bone repair material (P < 0.05). (4) In vivo micro-CT examination showed that the amount of new bone formation in the observation group was higher than that in the control group and the blank group    (P < 0.05), and that in the control group was higher than that in the blank group (P < 0.05). (5) In vivo hematoxylin-eosin staining and Masson staining showed that the bone repair effect of the observation group was better than that of the control and blank groups, and that in the control group was better than that in the blank group. (6) These findings confirmed that BMP-9 can further improve the bone induction properties of collagen-based bone repair materials. 

Key words: bone, material, collagen based bone, bone morphogenetic protein, bone defect, osteogenesis, osteoinduction

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