Inhibition of glycogen synthase kinase-3beta attenuates glucocorticoid-induced bone loss.
OBJECTIVE
Long-term glucocorticoid administration is known to induce bone deterioration. Glycogen synthase kinase-3beta (GSK-3beta) signaling reportedly participates in bone remodeling. This study investigated whether GSK-3beta inhibitor could regulate glucocorticoid-induced inhibition of osteoblast differentiation in vitro or bone mass in vivo.


METHODS
MC3T3-E1 osteoblasts were treated with kinase-inactive GSK-3beta mutant and 6-bromoindirubin-3'-oxim (BIO) and then exposed to 1microM dexamethasone. Survival and osteoblast differentiation of cell cultures were assessed by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-biotin nick end-labeling, quantitative RT-PCR, and von Kossa staining. Mineral density, biomechanical properties and microenvironments of BIO- and glucocorticoid-treated rat bone tissues were analyzed using dual-energy X-ray absorptiometry, material testing, and histomorphometry, respectively.


RESULTS
Glucocorticoid decreased levels of phosphorylated Ser9-GSK-3beta and beta-catenin in osteoblast cultures. Kinase-inactive GSK-3beta mutant and BIO treatments attenuated dexamethasone-induced inhibition of beta-catenin, Runx2 abundance, and osteoblast differentiation but suppressed glucocorticoid-induced apoptosis of cell cultures. Exogenous BIO treatment alleviated methylprednisolone-induced impairment of mineral density, biomechanical strength, trabecular bone volume, osteoblast surface, and bone formation rate of rat bone tissue. BIO treatment also attenuated glucocorticoid-induced promotion of osteoclast surface and marrow adipocyte volume in bone tissue. Bone cells adjacent to glucocorticoid-stressed bone tissue displayed strong phosphorylated Ser9-GSK-3beta and beta-catenin immunostaining following BIO treatment.


CONCLUSIONS
Inhibition of GSK-3beta abrogated glucocorticoid-induced bone loss by increasing beta-catenin- and Runx2-mediated osteoblast differentiation. Controlling GSK-3beta signaling in bone cells may be a strategy for preventing glucocorticoid-induced osteopenia.
Department of Medical Research, Chang Gung Memorial Hospital-Kaohsiung Medical Center, Taiwan. wangfs@ms33.hinet.net
Feng-Sheng
FS
Wang
Jih-Yang
JY
Ko
Lin-Hsiu
LH
Weng
Da-Wei
DW
Yeh
Huei-Jine
HJ
Ke
Shin-Long
SL
Wu
D016475:3T3 Cells; D000469:Alkaline Phosphatase / Q000378:metabolism; D000818:Animals; D017209:Apoptosis / Q000235:genetics / Q000502:physiology; D001696:Biomechanical Phenomena; D015153:Blotting, Western; D015519:Bone Density / Q000187:drug effects; D001857:Bone Matrix / Q000187:drug effects; D016723:Bone Remodeling / Q000187:drug effects*; D001842:Bone and Bones / Q000033:anatomy & histology / Q000187:drug effects; D002113:Calcification, Physiologic / Q000187:drug effects; D002454:Cell Differentiation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005938:Glucocorticoids / Q000037:antagonists & inhibitors* / Q000633:toxicity*; D038362:Glycogen Synthase Kinase 3 / Q000037:antagonists & inhibitors* / Q000235:genetics; D007150:Immunohistochemistry; D007211:Indoles / Q000494:pharmacology*; D008775:Methylprednisolone / Q000037:antagonists & inhibitors / Q000633:toxicity; D051379:Mice; D010006:Osteoblasts / Q000187:drug effects; D010091:Oximes / Q000494:pharmacology*; D051381:Rats; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection; D051176:beta Catenin / Q000378:metabolism
C483321:6-bromoindirubin-3'-oxime; D004791:Enzyme Inhibitors; D005938:Glucocorticoids; D007211:Indoles; D010091:Oximes; D051176:beta Catenin; D038362:Glycogen Synthase Kinase 3; D000469:Alkaline Phosphatase; D008775:Methylprednisolone