Expression profiling of human adipose tissue derived stem cells cultured at various oxygen tensions

Adipose tissue is a plentiful and easily accessible source of mesenchymal stem cells that have been shown to be multi potent and possibly have immuno suppressive capacity(Zuk, Zhu et al. 2002; McIntosh, Zvonic et al. 2006; Prichard, Reichert et al. 2008). Several studies have identified hypoxia and the molecular effector of reduced oxygen tension, HIF1α to be essential in the differentiation processes, stem cell proliferation and survival as well as in maintenance of stem cell characteristics (Lennon, Edmison et al. 2001; Goda, Ryan et al. 2003; Wang, Fermor et al. 2005; Lin, Lee et al. 2006; Malladi, Xu et al. 2007; Ohnishi, Yasuda et al. 2007). The fact that oxygen has a central role in cell development and metabolism makes the regulation of oxygen tension a valuable tool in the attempt of exploiting stem cells to their full potential. In the current study we investigate the transcriptional changes caused by two weeks of monolayer hypoxic expansion of human adipose tissue derived stem cells (ASCs). Application of the XVivo hypoxic workstation (BioSpherix, Redfield, NY) made it possible to conduct the expansion of six ASC lines at four different hypoxic conditions in parallel covering the range from mild (15% oxygen) to full hypoxic (1% oxygen) conditions. In this comprehensive investigation the Illumina bead microarray platform was employed to give a profound and new insight in the molecular changes effectuated by hypoxic expansion. The findings of this study demonstrate the significance of biologic variation in the transcriptional response to mild hypoxic expansion and display evidence that full hypoxic growth conditions at 1% oxygen tension selects for a more prolific cell population of reduced chondrogenic potential. Keywords: hypoxia response To investigate the transcriptional response to hypoxia, adipose tissue derived stem cells (ASCs) from six female individuals were cultured for 14 days at different oxygen tensions (ambient, 15%, 10%, 5% and 1%). Gene expression levels at each oxygen tension were compared to ambient conditions (Ambient oxygen tension) using Illumina Human-6 v2.0 microarrays.

脂肪组织是一类来源丰富且易于获取的间充质干细胞（mesenchymal stem cells）库，该类细胞已被证实具有多向分化潜能，且可能具备免疫抑制能力（Zuk、Zhu等，2002；McIntosh、Zvonic等，2006；Prichard、Reichert等，2008）。多项研究表明，缺氧以及氧分压降低的核心分子效应物缺氧诱导因子-1α（hypoxia-inducible factor 1α, HIF1α），在干细胞分化、增殖与存活以及干细胞特性维持过程中发挥关键调控作用（Lennon、Edmison等，2001；Goda、Ryan等，2003；Wang、Fermor等，2005；Lin、Lee等，2006；Malladi、Xu等，2007；Ohnishi、Yasuda等，2007）。鉴于氧在细胞发育与代谢中具有核心地位，氧分压调控成为充分挖掘干细胞应用潜力的重要技术手段。
本研究针对人类脂肪来源干细胞（adipose tissue-derived stem cells, ASCs）开展为期两周的单层贴壁缺氧扩增，探究该过程引发的转录组变化。本研究采用XVivo缺氧培养工作站（BioSpherix公司，纽约州雷德菲尔德），实现了6株ASC系在4种不同缺氧条件下的平行扩增，覆盖了从轻度缺氧（15%氧浓度）到重度缺氧（1%氧浓度）的全范围氧分压梯度。本研究采用Illumina微珠芯片检测平台开展全面分析，以期为缺氧扩增引发的分子调控变化提供全新的深度认知。
本研究结果证实，轻度缺氧扩增过程中的转录应答存在显著的生物学个体差异；同时发现，1%氧浓度的重度缺氧培养条件可筛选出增殖能力更强但成软骨分化潜能降低的细胞群体。
关键词：缺氧应答
为探究缺氧应答的转录调控效应，本研究从6名女性个体体内分离得到脂肪来源干细胞（ASCs），在不同氧分压条件（常氧、15%、10%、5%及1%氧浓度）下培养14天。采用Illumina Human-6 v2.0基因芯片，将各氧分压条件下的基因表达水平与常氧组（常氧分压）进行对比分析。