A mineralization-associated membrane protein plays a role in the biological functions of the peptide-coated bovine hydroxyapatite.
OBJECTIVE
Anorganic bovine mineral coated with a cell-binding peptide (P-15) is superior to anorganic bovine mineral alone in the treatment of periodontal osseous defects. However, the molecular interactions between P-15 and periodontal ligament fibroblasts remain unclear.


METHODS
We first compared the in vitro osteogenic activities between periodontal ligament fibroblasts cultured with anorganic bovine mineral alone and with the P-15/anorganic bovine mineral combination. We then harvested the periodontal ligament cell lysate, incubated it with various graft materials, and then washed it to remove unbound proteins. The bound proteins were eluted from graft materials and analyzed using electrophoresis, followed by mass spectrometry and then western blotting. Finally, a neutralizing antibody against one bound protein was added to the cell cultures to repeat the osteogenic assays to clarify its role in the in vitro effects of the P-15/anorganic bovine mineral combination.


RESULTS
Cells treated with P-15/anorganic bovine mineral were more viable and showed greater osteogenic activities than cells treated with anorganic bovine mineral alone and the no-graft control. Annexin II, a mineralization-associated protein, bound to P-15/anorganic bovine mineral significantly more than to anorganic bovine mineral alone. The addition of neutralizing antibody for annexin II decreased the osteogenic activities of the P-15/anorganic bovine mineral combination.


CONCLUSIONS
Annexin II of periodontal ligament fibroblasts interacted with the peptide of P-15, and was partially responsible for better in vitro osteogenesis in the P-15 graft.
Yuan
K
K
Institute of Oral Medicine and Department of Biochemistry, Medical School, National Cheng Kung University, and Department of Dentistry, Tainan Municipal Hospital, Tainan, Taiwan. kuoyuan@mail.ncku.edu.tw
Huang
J-S
JS
Hsu
C-W
CW
Hung
I-J
IJ
D000328:Adult; D000469:Alkaline Phosphatase / Q000032:analysis; D017306:Annexin A2 / Q000378:metabolism* / Q000502:physiology; D000880:Anthraquinones; D015153:Blotting, Western; D001857:Bone Matrix / Q000378:metabolism / Q000637:transplantation*; D018786:Bone Substitutes / Q000627:therapeutic use*; D002113:Calcification, Physiologic / Q000187:drug effects / Q000502:physiology; D002470:Cell Survival; D002478:Cells, Cultured; D004396:Coloring Agents; D017077:Culture Media, Conditioned; D017886:Durapatite / Q000627:therapeutic use*; D004591:Electrophoresis, Polyacrylamide Gel; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D008297:Male; D013058:Mass Spectrometry; D010012:Osteogenesis / Q000187:drug effects / Q000502:physiology*; D010513:Periodontal Ligament / Q000187:drug effects / Q000378:metabolism; D011485:Protein Binding
D017306:Annexin A2; D000880:Anthraquinones; D018786:Bone Substitutes; D004396:Coloring Agents; D017077:Culture Media, Conditioned; C118792:OsteoGraf-N; C403137:PepGen P-15; C010078:alizarin; D017886:Durapatite; D000469:Alkaline Phosphatase