Combined effects of melatonin and FGF-2 on mouse preosteoblast behavior within interconnected porous hydroxyapatite ceramics - in vitro analysis.
Objective Biocompatible materials such as interconnected porous hydroxyapatite ceramics (IP-CHA) loaded with osteogenic cells and bioactive agents are part of an evolving concept for overcoming craniofacial defects by use of artificial bone tissue regeneration. Amongst the bioactive agents, melatonin (MEL) and basic fibroblast growth factor (FGF-2) have been independently reported to induce osteoblastic activity. The present in vitro study was undertaken to examine the relationship between these two bioactive agents and their combinatory effects on osteoblastic activity and mineralization in vitro. Material and Methods Mouse preosteoblast cells (MC3T3-E1) were seeded and cultured within cylindrical type of IP-CHA block (ø 4x7 mm) by vacuum-assisted method. The IP-CHA/MC3T3 composites were subjected to FGF-2 and/or MEL. The proliferation assay, alkaline phosphatase enzyme activity (ALP), mRNA expressions of late bone markers, namely Osteocalcin (OCN) and Osteopontin (OPN), and Alizarin Red staining were examined over a period of 7 days. Results FGF-2 mainly enhanced the proliferation of MC3T3-E1 cells within the IP-CHA constructs. MEL mainly induced the mRNA expression of late bone markers (OCN and OPN) and showed increased ALP activity of MC3T3 cells cultured within IP-CHA construct. Moreover, the combination of FGF-2 and MEL showed increased osteogenic activity within the IP-CHA construct in terms of cell proliferation, upregulated expressions of OCN and OPN, increased ALP activity and mineralization with Alizarin Red. The synergy of the proliferative potential of FGF-2 and the differentiation potential of MEL showed increased osteogenic activity in MC3T3-E1 cells cultured within IP-CHA constructs. Conclusion These findings indicate that the combination of FGF-2 and MEL may be utilized with biocompatible materials to attain augmented osteogenic activity and mineralization.
Department of Oral and Maxillofacial Surgery, Institute of Biomedical and Health Sciences, Hiroshima University, Hiroshima, Japan.
Mohammad Zeshaan
MZ
Rahman
Department of Oral and Maxillofacial Surgery, Institute of Biomedical and Health Sciences, Hiroshima University, Hiroshima, Japan.
Hideo
H
Shigeishi
Department of Oral and Maxillofacial Surgery, Institute of Biomedical and Health Sciences, Hiroshima University, Hiroshima, Japan.
Kazuki
K
Sasaki
Department of Oral and Maxillofacial Surgery, Institute of Biomedical and Health Sciences, Hiroshima University, Hiroshima, Japan.
Akira
A
Ota
Department of Oral and Maxillofacial Surgery, Institute of Biomedical and Health Sciences, Hiroshima University, Hiroshima, Japan.
Kouji
K
Ohta
Department of Oral and Maxillofacial Surgery, Institute of Biomedical and Health Sciences, Hiroshima University, Hiroshima, Japan.
Masaaki
M
Takechi
D000469:Alkaline Phosphatase / Q000032:analysis; D000818:Animals; D018786:Bone Substitutes / Q000494:pharmacology*; D002113:Calcification, Physiologic / Q000187:drug effects; D002454:Cell Differentiation / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D017886:Durapatite / Q000494:pharmacology*; D016222:Fibroblast Growth Factor 2 / Q000494:pharmacology*; D008422:Materials Testing; D008550:Melatonin / Q000494:pharmacology*; D051379:Mice; D008855:Microscopy, Electron, Scanning; D010006:Osteoblasts / Q000187:drug effects*; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013997:Time Factors
D018786:Bone Substitutes; D016222:Fibroblast Growth Factor 2; D017886:Durapatite; D000469:Alkaline Phosphatase; D008550:Melatonin