title: In situ quasi-static and dynamic nanoindentation tests on calcified nodules formed by osteoblasts: Implication of glucocorticoids responsible for osteoblast calcification.
abstract: The functional requirements of regenerated calcified tissues are that they enable the tissues to bear a variety of imposed stress and consequent contact-induced strain without substantial fracture. Here we demonstrate the effects of glucocorticoid hormones such as dexamethasone and hydrocortisone on the nanomechanical properties of calcified nodules formed by mouse osteoblastic MC3T3-E1 cells in differentiation-inducing medium containing ascorbic acid and β-glycerophosphate. Neither cell proliferation nor calcium deposition, evaluated using alizarin red and von Kossa staining, was affected by dexamethasone. On the other hand, calcified nodules formed in the presence of dexamethasone were significantly harder and stiffer than those formed in their absence. In particular, a series of nanoindentation tests revealed that the calcified nodules formed in the presence of dexamethasone showed enhanced stiffness against dynamic strain as compared to a quasi-static load. Furthermore, Raman spectroscopy revealed that dexamethasone and hydrocortisone increased the apatite/matrix ratio and lowered that of carbonate in the nodules. Our results suggest that glucocorticoids are required for in vitro formation by osteoblasts of more mature calcified nodules containing apatite/phosphate.
authors.affiliation: Department of Biochemistry, Showa University School of Dentistry, Tokyo, Japan; Department of Orthodontics, Showa University School of Dentistry, Tokyo, Japan.
authors.forename: Sho
authors.initials: S
authors.lastname: Miyamoto
authors.affiliation: Department of Biochemistry, Showa University School of Dentistry, Tokyo, Japan. Electronic address: yoichim@dent.showa-u.ac.jp.
authors.forename: Yoichi
authors.initials: Y
authors.lastname: Miyamoto
authors.affiliation: Department of Conservative Dentistry, Division of Biomaterials and Engineering, Showa University School of Dentistry, Tokyo, Japan.
authors.forename: Yo
authors.initials: Y
authors.lastname: Shibata
authors.affiliation: Department of Biochemistry, Showa University School of Dentistry, Tokyo, Japan.
authors.forename: Kentaro
authors.initials: K
authors.lastname: Yoshimura
authors.affiliation: Department of Biochemistry, Showa University School of Dentistry, Tokyo, Japan; Department of Orthodontics, Showa University School of Dentistry, Tokyo, Japan.
authors.forename: Eri
authors.initials: E
authors.lastname: Izumida
authors.affiliation: Department of Biochemistry, Showa University School of Dentistry, Tokyo, Japan.
authors.forename: Hiroaki
authors.initials: H
authors.lastname: Suzuki
authors.affiliation: Department of Conservative Dentistry, Division of Biomaterials and Engineering, Showa University School of Dentistry, Tokyo, Japan.
authors.forename: Takashi
authors.initials: T
authors.lastname: Miyazaki
authors.affiliation: Department of Orthodontics, Showa University School of Dentistry, Tokyo, Japan.
authors.forename: Koutaro
authors.initials: K
authors.lastname: Maki
authors.affiliation: Department of Biochemistry, Showa University School of Dentistry, Tokyo, Japan.
authors.forename: Ryutaro
authors.initials: R
authors.lastname: Kamijo
mesh_terms: D016475:3T3 Cells; D000818:Animals; D002113:Calcification, Physiologic; D005938:Glucocorticoids / Q000494:pharmacology*; D051379:Mice; D010006:Osteoblasts / Q000187:drug effects / Q000473:pathology*
chemical_list: D005938:Glucocorticoids
keywords: Calcification; Hardness; Hydroxyapatite; Nanoindentation; Osteoblast