Mapping the osteocytic cell response to fluid flow using RNA-Seq
Mapping the osteocytic cell response to fluid flow using RNA-Seq
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DOI：
10.1016/j.jbiomech.2015.10.045
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发表时间：
2015-12-16
影响因子：
2.4
通讯作者：
Donahue, Henry J.
中科院分区：
文献类型：
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作者：
Govey, Peter M.;Kawasawa, Yuka Imamura;Donahue, Henry J.
骨对机械负荷的适应性通过机械敏感骨细胞的信号转导来调节。矿物质包埋的骨细胞经历应变诱导的间质液流动和流体剪切应力，并且基因表达的广泛变化是调节骨转换的信号通路中的关键组分。RNA测序分析，或RNA-Seq，能够比以前更完整地表征机械响应转录组调控。我们假设骨细胞MLO-Y 4细胞的RNA-Seq揭示了先前流体流动并使用基因微阵列分析的细胞中预期的和新的基因转录调控。使MLO-Y 4细胞在1 Pa振荡流体剪切应力下流动2小时，并后孵育2小时。原始样品的RNA-Seq检测到55个流体流动调节基因转录物（p校正1.5倍或降低< 0.67倍，而55个中的10个符合微阵列的该截止值。分析在调控模式上是互补的，尽管只有6个转录本在RNA-Seq和微阵列分析中是显著的：Cxcl 5、Oxcl 1、Zc 3 h12 a、Ereg、Slc 2a 1和Egln 1。作为基因本体分析推断的广泛炎症反应的一部分，我们再次观察到炎症C-X-C基序趋化因子的最大上调，以及新涉及的HIF-1 α和AMPK信号通路。重要的是，我们检测了预期的流体流动敏感性转录物（例如Nos 2 [iNOS]、Prgs 2 [考克斯-2]、Ccl 7）和先前未鉴定为流动敏感性的转录物，例如Ccl 2。我们发现RNA-Seq优于微阵列，因为它具有更大的动态范围和分析基因表达无偏估计的能力，为我们了解骨细胞信号传导提供了信息。(C)2015爱思唯尔有限公司版权所有。
Bone adaptation to mechanical loading is regulated via signal transduction by mechano-sensing osteocytes. Mineral-embedded osteocytes experience strain-induced interstitial fluid flow and fluid shear stress, and broad shifts in gene expression are key components in the signaling pathways that regulate bone turnover. RNA sequencing analysis, or RNA-Seq, enables more complete characterization of mechano-responsive transcriptome regulation than previously possible. We hypothesized that RNA-Seq of osteocytic MLO-Y4 cells reveals both expected and novel gene transcript regulation in cells previously fluid flowed and analyzed using gene microarrays. MLO-Y4 cells were flowed for 2 h with 1 Pa oscillating fluid shear stress and post-incubated 2 h. RNA-Seq of original samples detected 55 fluid flow-regulated gene transcripts (p-corrected 1.5-fold or decreased < 0.67-fold whereas 10 of 55 met this cut-off by microarray. Analyses were complimentary in patterns of regulation, though only 6 transcripts were significant in both RNA-Seq and microarray analyses: Cxcl5, Oxcl1, Zc3h12a, Ereg, Slc2a1, and Egln1. As part of a broad inflammatory response inferred by gene ontology analyses, we again observed greatest up-regulation of inflammatory C-X-C motif chemokines, and newly implicated HIF-1 alpha and AMPK signaling pathways. Importantly, we detected both expected fluid flow-sensitive transcripts (e.g. Nos2 [iNOS], Prgs2 [COX-2], Ccl7) and transcripts not previously identified as flow-sensitive, e.g. Ccl2. We found RNA-Seq advantageous over microarrays because of its greater dynamic range and ability to analyze unbiased estimation of gene expression, informing our understanding of osteocyte signaling. (C) 2015 Elsevier Ltd. All rights reserved.