Fluid shear stress promotes osteoblast proliferation via the Gαq-ERK5 signaling pathway
Fluid shear stress promotes osteoblast proliferation via the Gαq-ERK5 signaling pathway
复制标题
流体剪切应力通过 G α q-ERK5 信号通路促进成骨细胞增殖
DOI：
10.1080/03008207.2016.1181063
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发表时间：
2016-01-01
影响因子：
2.9
通讯作者：
Xia Yayi
中科院分区：
文献类型：
--
作者：
Zhang Bo;Geng Bin;Xia Yayi
流体剪应力(FSS)是一种普遍存在的机械刺激，能有效地促进成骨细胞的增殖。此前，我们报道了细胞外信号调节蛋白5(ERK5)在FSS诱导的成骨细胞增殖中起着至关重要的作用。然而，FSS通过ERK5激活促进成骨细胞增殖的确切机制尚不清楚。本研究的目的是确定GαQ在FSS诱导的ERK5磷酸化和成骨细胞增殖中的关键作用，以及GαQ-ERK5通路的下游靶点。将50 nM GαQ siRNA导入MC3T3-E1细胞，用5 mM的XMD8-92(一种高选择性的ERK5活性抑制剂)和/或FSS(12dyn/cm(2))处理。用四甲基偶氮唑蓝(3-(4，5-dimethylthiazol-2-yl)-2，5-diphenyltetrazolium)比色法检测细胞增殖。Western印迹分析GαQ、P-ERK5、ERK5、Cyclin B1、CDK1的蛋白表达水平。生理性FSS暴露60min可显著促进MC3T3-E1细胞的增殖，但这种作用可被siRNA介导的GαQ基因敲除或XMD8-92抑制ERK5活性所抑制，提示GαQ和ERK5可能参与了FSS促进的成骨细胞增殖。此外，GαQ siRNA显著抑制ERK5的磷酸化。此外，我们的研究还发现，FSS处理MC3T3-E1细胞60min后，Cyclin B1和CDK1的蛋白表达水平显著上调，这种上调可被GαQ siRNA或XMD8-92显著阻断。我们认为FSS作用于GαQ-ERK5信号通路，上调Cyclin B1和CDK1的表达，从而导致MC3T3-E1细胞的增殖。因此，GαQ-ERK5信号通路可能为骨代谢疾病的治疗提供有用的信息。
Fluid shear stress (FSS) is a ubiquitous mechanical stimulus that potently promotes osteoblast proliferation. Previously, we reported that extracellular signal-regulated kinase 5 (ERK5) is essential for FSS-induced osteoblast proliferation. However, the precise mechanism by which FSS promotes osteoblast proliferation via ERK5 activation is poorly understood. The aim of this study was to determine the critical role of G alpha q in FSS-induced ERK5 phosphorylation and osteoblast proliferation, as well as the downstream targets of the G alpha q-ERK5 pathway. MC3T3-E1 cells were transfected with 50 nM G alpha q siRNA, treated with 5 mM XMD8-92 (a highly selective inhibitor of ERK5 activity), and/or exposed to FSS (12 dyn/cm(2)). Cell proliferation was evaluated using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The protein expression levels of G alpha q, P-ERK5, ERK5, Cyclin B1, and CDK1 were analyzed by Western blot. Physiological FSS exposure for 60 min remarkably promoted MC3T3-E1 cell proliferation, however, this effect was suppressed by siRNA-mediated G alpha q knockdown or inhibition of ERK5 activity by XMD8-92 treatment, suggesting that G alpha q and ERK5 might modulate FSS-increased osteoblast proliferation. Furthermore, ERK5 phosphorylation was dramatically inhibited by G alpha q siRNA. In addition, our study further revealed that FSS treatment of MC3T3-E1 cells for 60 min markedly upregulated the protein expression levels of Cyclin B1 and CDK1, and this increased expression was predominantly blocked by G alpha q siRNA or XMD8-92 treatment. We propose that FSS acts on the G alpha q-ERK5 signaling pathway to upregulate Cyclin B1 and CDK1 expression, thereby resulting in MC3T3-E1 cell proliferation. Thus, the G alpha q-ERK5 signaling pathway may provide useful information regarding the treatment of bone metabolic disease.