The PI3K/Akt/FOXO3a/p27Kip1 signaling contributes to anti-inflammatory drug-suppressed proliferation of human osteoblasts

Akt has been reported to suppress p27(Kip1) promoter activity through Forkhead box O (FOXO) in different kinds of cells. Previous studies indicated that anti-inflammatory drugs up-regulated p27(Kip1), and this effect might play an important role in anti-inflammatory drug-induced cell cycle arrest of human osteoblasts (hOBs). In this study, we hypothesized that these drugs might increase p27(Kip1) expression in hOBs by altering the Akt/FOXO signaling. We tested this hypothesis by examining the influences of three anti-inflammatory drugs on the levels and/or activities of Akt, FOXO and p27(Kip1) as well as the relationship between these factors and proliferation of hOBs. We tested the effects of indomethacin (10(-5) and 10(-4) M), celecoxib(10(-6) and 10(-5) M), and dexamethasone (10(-7) and 10(-6) M) using PI3K inhibitor, LY294002 (10(-5) M) as the basis of comparison. The three drugs suppressed the canonical level of phosphorylated Akt in hOBs. This was accompanied by elevated FOXO3a level and increased promoter activity, mRNA expression and protein level of p27(Kip1). Furthermore, the anti-inflammatory drugs suppressed the EGF-induced increases in proliferation, phosphorylation, and nucleus translocation of Akt. Simultaneously, they suppressed EGF-induced decreases of FOXO3a nucleus accumulation and p27(Kip1) mRNA expression. On the other hand, FOXO silencing significantly attenuated the drug-induced up-regulation of p27(Kip1) and suppression of proliferation in hOBs. To the best of our knowledge, this study represents the first to demonstrate that Akt/FOXO3a/p27(Kip1) pathway contributes to suppression of hOB proliferation by anti-inflammatory drugs. We suggest that anti-inflammatory drugs suppress hOB proliferation, at least partly, through inactivating Akt, activating FOXO3a, and eventually up-regulating p27(Kip1) expression. (C) 2009 Elsevier Inc. All rights reserved.