Zhang Hongyu, Li Li, Dong Qian, Wang Yufeng, Feng Qiaoling, Ou Xinying, Zhou Pengfei, He Tongchuan, Luo Jinyong
Cell Physiol Biochem. 2015;37(2):548-62. doi: 10.1159/000430376.
BACKGROUND/AIMS: BMP9 is highly capable of promoting osteogenic differentiation of mesenchymal stem cells (MSCs) although the molecular mechanism involved is largely unknown. Here, we explored the detail role of PKA/CREB signaling in BMP9-induced osteogenic differentiation.
Activation status of PKA/CREB signaling is assessed by nonradioactive assay and Western blot. Using PKA inhibitors and a dominant negative protein of CREB (A-CREB), we investigated the effect of PKA/CREB signaling on BMP9-induced osteogenic differentiation.
We found that BMP9 promotes PKA activity and enhances CREB phosphorylation in MSCs. BMP9 is shown to down-regulate protein kinase A inhibitor γ (PKIγ) expression. We demonstrated that PKA inhibitors suppress BMP9-induced early osteogenic marker alkaline phosphatase (ALP) activity in MSCs as well as late osteogenic markers osteopontin (OPN), osteocalcin (OCN) and matrix mineralization. We found that PKA inhibitor reduces BMP9-induced Runx2 activation and p38 phosphorylation in MSCs. Lastly, interference of CREB function by A-CREB decreased BMP9-induced osteogenic differentiation as well.
Our results revealed that BMP9 may activate PKA/CREB signaling in MSCs through suppression of PKIγ expression. It is noteworthy that inhibition of PKA/CREB signaling may impair BMP9-induced osteogenic differentiation of MSCs, implying that activation of PKA/CREB signaling is required for BMP9 osteoinductive activity.
背景/目的：骨形态发生蛋白9（BMP9）具有很强的促进间充质干细胞（MSC）成骨分化的能力，但其涉及的分子机制在很大程度上尚不清楚。在此，我们探讨了蛋白激酶A（PKA）/环磷腺苷效应元件结合蛋白（CREB）信号通路在BMP9诱导的成骨分化中的具体作用。
通过非放射性检测和蛋白质印迹法评估PKA/CREB信号通路的激活状态。使用PKA抑制剂和CREB的显性负性蛋白（A-CREB），我们研究了PKA/CREB信号通路对BMP9诱导的成骨分化的影响。
我们发现BMP9可促进MSC中的PKA活性并增强CREB磷酸化。BMP9可下调蛋白激酶A抑制剂γ（PKIγ）的表达。我们证明PKA抑制剂可抑制BMP9诱导的MSC早期成骨标志物碱性磷酸酶（ALP）活性以及晚期成骨标志物骨桥蛋白（OPN）、骨钙素（OCN）和基质矿化。我们发现PKA抑制剂可降低BMP9诱导的MSC中Runx2激活和p38磷酸化。最后，A-CREB对CREB功能的干扰也降低了BMP9诱导的成骨分化。
我们的结果表明，BMP9可能通过抑制PKIγ表达激活MSC中的PKA/CREB信号通路。值得注意的是，抑制PKA/CREB信号通路可能会损害BMP9诱导的MSC成骨分化，这意味着激活PKA/CREB信号通路是BMP9骨诱导活性所必需的。
