In vitro assay of mineralized-tissue formation on titanium using fluorescent staining with calcein blue.
Many studies have examined mineralized-tissue formation on titanium in vivo and in vitro; however, no precise method for measuring the mineralized tissue produced by cultured osteoblastic cells on titanium in vitro has been established. This study developed a method for measuring mineralized-tissue formation by cultured rat osteoblastic cells on titanium in vitro, and re-evaluated the effects of modifying the titanium surface. We used calcein blue, which binds to mineralized tissue, and measured the resulting fluorescence under ultraviolet light. A 1-h incubation with 3.1x10(-3)M calcein blue resulted in sufficient fluorescence of bone-like nodules on the titanium. Consequently, we found that osteoblastic cells produced larger bone-like nodules on titanium than in culture dishes. Fewer bone-like nodules formed on calcium-ion impregnated titanium than on pure titanium. However, when bisphosphonate was immobilized on the calcium-ion impregnated titanium, more bone-like nodules formed than on pure titanium. The results suggest that bisphosphonate immobilization on titanium is useful for stimulating mineralized-tissue formation on titanium implants.
Goto
Tetsuya
T
Department of Oral Anatomy, Kyushu Dental College, Kitakyushu 803-8580, Japan. tgoto@kyu-dent.ac.jp
Kajiwara
Hiroshi
H
Yoshinari
Masao
M
Fukuhara
Eiji
E
Kobayashi
Shigeru
S
Tanaka
Teruo
T
D000818:Animals; D002118:Calcium / Q000378:metabolism; D002478:Cells, Cultured; D004164:Diphosphonates / Q000737:chemistry; D005452:Fluoresceins / Q000494:pharmacology*; D005456:Fluorescent Dyes / Q000494:pharmacology; D006801:Humans; D066298:In Vitro Techniques; D007477:Ions; D008856:Microscopy, Fluorescence / Q000379:methods*; D010006:Osteoblasts / Q000378:metabolism; D011485:Protein Binding; D051381:Rats; D017208:Rats, Wistar; D013997:Time Factors; D014025:Titanium / Q000737:chemistry*
D004164:Diphosphonates; D005452:Fluoresceins; D005456:Fluorescent Dyes; D007477:Ions; D014025:Titanium; D002118:Calcium; C007740:fluorexon