Glucocorticoids inhibit cell cycle progression in differentiating osteoblasts via glycogen synthase kinase-3β

Abstract

Differentiating osteoblasts in culture undergo a commitment stage, during which cobblestone-like cells grow to high density past confluency. In contrast to earlier proliferative stages, the cell cycle during this commitment stage is inhibited by glucocorticoids (GC). Chronic GC treatment also impedes mineral deposition if steroid administration commences early enough during commitment. This study defines a role for glycogen synthase kinase-3β (GSK3β) and its target, c-Myc, in the GC-sensitive osteoblast persistent cell cycle. c-Myc levels decreased as cells reached confluence, but then increased during growth to high density. GC administration at this stage resulted in down-regulation of c-Myc. This was accompanied by GC-mediated attenuation of GSK3β Ser9 inhibitory phosphorylation and increased GSK3β kinase activity. Down-regulation of c-Myc was attributable to enhanced Thr58 phosphorylation, leading to accelerated degradation. In contrast, GC did not inhibit the c-Myc synthesis rate or the level of β-catenin, a transcriptional coactivator of c-myc. The attenuated cell cycle and the reduced c-Myc level were returned to control levels by specific inhibition of GSK3β using lithium chloride. These results suggest that tonal GSK3β repression at the cobblestone stage of osteoblast differentiation permits osteoblast growth to high density. GC interfere with this growth-permissive axis by GSK3β activation, resulting in c-Myc down-regulation and impediment of the G1/S cell cycle transition.