J Korean Assoc Oral Maxillofac Surg.  2021 Dec;47(6):454-464. 10.5125/jkaoms.2021.47.6.454.

Effects of the combination of bone morphogenetic protein-2 and nano-hydroxyapatite on the osseointegration of dental implants

Affiliations
  • 1Department of Dentistry, Oral and Maxillofacial Surgery, Seoul National University Dental Hospital, Seoul, Korea
  • 2Department of Medical Biotechnology, College of Life Science and Biotechnology, Dongguk University, Seoul, Korea
  • 3Department of Oral and Maxillofacial Surgery, School of Dentistry, Seoul National University, Seoul, Korea
  • 4Dental Life Science Research Institute and Clinical Translational Research Center for Dental Science, Seoul National University Dental Hospital, Seoul, Korea

Abstract


Objectives
This study aimed to investigate the in vitro osteoinductivity of the combination of bone morphogenetic protein-2 (BMP-2) and nanohydroxyapatite (nHAp) and the in vivo effects of implants coated with nHAp/BMP-2.
Materials and Methods
To evaluate the in vitro efficacy of nHAp/BMP-2 on bone formation, bone marrow-derived mesenchymal stem cells (BMMSCs) were seeded onto titanium disks coated with collagen (Col), Col/nHAp, or Col/nHAp/BMP-2. Protein levels were determined by a biochemical assay and reverse transcriptase-polymerase chain reaction. Stem cell differentiation was analyzed by flow cytometry. For in vivo studies with mice, Col, Col/nHAp, and Col/nHAp/BMP-2 were injected in subcutaneous pockets. Titanium implants or implants coated with Col/nHAp/BMP-2 were placed bilaterally on rabbit tibias and evaluated for 4 weeks.
Results
In the in vitro study, BM-MSCs on Col/nHAp/BMP-2 showed reduced levels of CD73, CD90, and CD105 and increased levels of glycos-aminoglycan, osteopontin, and alkaline phosphatase activity. After 4 weeks, the Col/nHAp/BMP-2 implant showed greater bone formation than the control (P=0.07), while no differences were observed in bone implant contact and removal torque.
Conclusion
These results suggest that a combination of BMP-2 and an nHAp carrier would activate osseointegration on dental implant surfaces.

Keyword

Bone morphogenetic protein 2; Hydroxyapatite; Collagen; Dental implant; Osseointegration

Figure

  • Fig. 1 Topographic and three-dimensional atomic force microscopic images of the coatings. Titanium: untreated titanium disk, Col: collagen-coated titanium surface, Col/BMP-2: collagen/bone morphogenetic protein-2 (BMP-2)-coated titanium surface, Col/nHAp/BMP-2: collagen/nano-hydroxyapatite/BMP-2-coated titanium surface. (RMS: root mean square)

  • Fig. 2 Static water contact angle on titanium surface. A. Negative control (resorbable blasted media surface titanium). B. Collagen-coated surface. C. Collagen/BMP-2-coated surface. D. Collagen/nHAp/BMP-2-coated surface. (BMP-2: bone morphogenetic protein-2, nHAp: nano-hydroxyapatite)

  • Fig. 3 In vitro release curve of bone morphogenetic protein-2 (BMP-2) from various surface-treated titanium disks. The results are shown as mean±standard deviation values (n=3). An increase in BMP-2 release was observed within 24 hours. A sustained release in the Col/nHAp/BMP-2 group was observed between 1-5 days. (Col: collagen, nHAp: nano-hydroxyapatite)

  • Fig. 4 Bone marrow-derived mesenchymal stem cell (BM-MSC) differentiation on differently treated titanium disks. A. Flow cytometry of typical CD markers present on BM-MSCs cultured on the Col-, Col/nHAp-, or Col/nHAp/BMP-2-coated titanium disks. CD73+, CD90+, and CD105+ cells were observed prior to differentiation. Reduction in mesenchymal CD73+, CD90+, and CD105+ cells was observed in the Col/nHAp/BMP-2 group. B. Biochemical assays for alkaline phosphatase (ALP) activity and intracellular glycosaminoglycan (GAG) and osteopontin expression in BM-MSCs. Cells on Col/nHAp/BMP-2 exhibited the highest expression. C. mRNA expression of different osteoblastic markers in BM-MSC. The highest expression was observed for type III collagen (Col III), osteocalcin, and osteoprotegerin in cells grown on Col/nHAp/BMP-2. The levels of type I collagen (Col I), osteopontin, osteonectin, and BMP-2 expression were higher in BM-MSCs on Col/nHAp and Col/nHAp/BMP-2 than in those with Col enrichment. (Col: collagen, nHAp: nano-hydroxyapatite, BMP-2: bone morphogenetic protein-2)

  • Fig. 5 Histological images of the mouse subcutaneous pocket injection model. Col (A, D, G, J, M, P), Col/nHAp (B, E, H, K, N, Q), and Col/nHAp/BMP (C, F, I, L, O, R). Macroscopic images (A-C), H&E staining (D-F), Masson’s trichrome (MT) staining (G-I), von Kossa staining (J-L), osteonectin staining (M-O), and CD31 staining (P-R) (D-O: ×100, P-R: ×200, scale bars=200 µm). (Col: collagen, nHAp: nano-hydroxyapatite, BMP-2: bone morphogenetic protein-2)

  • Fig. 6 Implant installation on rabbit tibia. A. Photograph of titanium implant (left) and Col/nHAp/BMP-2-coated implant (right). B-D. Bone implant contact, new bone area, and removal torque. At 4 weeks after implant installation on rabbit tibia, Col/nHAp/BMP-2 exhibited increased new bone area compared with the negative control (P=0.07), whereas bone implant contact and removal torque exhibited no significant difference, although the mean values were higher. (Col: collagen, nHAp: nano-hydroxyapatite, BMP-2: bone morphogenetic protein-2)


Reference

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