Identifying pH independent hypoxia induced genes in human squamous cell carcinomas in vitro
Identifying pH independent hypoxia induced genes in human squamous cell carcinomas in vitro
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DOI：
10.3109/02841861003614343
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发表时间：
2010-10-01
期刊：
影响因子：
3.1
通讯作者：
Alsner, Jan
中科院分区：
文献类型：
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作者：
Sorensen, Brita Singers;Toustrup, Kasper;Alsner, Jan
背景资料。低氧上调的基因被认为是肿瘤缺氧的内源性标志物。然而，大多数被调查的基因都显示出不一致的结果，这导致了人们对它们是否有能力成为真正的缺氧制造者的担忧。以往的研究表明，细胞外pH(PH E)可以影响低氧诱导基因的表达。方法：研究方法。将5种不同的人细胞系(SiHA、FaDu(DD)、UTSCC5、UTSCC14和UTSCC15)暴露于不同的氧气浓度和pH值(7.5或6.3)中，并用基因芯片(Affymetrix-Human Genome U133 Plus 2.0阵列)分析基因表达。结果。使用SAM对其中两个细胞系进行分析，确定了461个能够将四组细胞区分开来的试管，这些试管能够将四个组分开：正常氧，正常pH，低氧，正常pH，“正常氧，低pH”和“低氧，低pH”。从这两个细胞系中可以鉴定出在低氧条件下诱导的与pH无关的一组分，这些组分包括HIG2、NDRG1、PAI1和RORA。定量聚合酶链式反应的进一步验证强调了使用更多的细胞系来获得稳健的基因表达谱的必要性。为了在更多的细胞系中特异性地筛选pH非依赖性的低氧调节基因，我们分析了FaDu(DD)、UTSCC5、UTSCC14和UTSCC15的数据，以确定每个细胞系中受低氧诱导的基因，其中这种诱导不受低pH的影响，并且该基因不受低pH单独诱导的显著影响。每个细胞系有65-122个符合这些标准的试剂组。对于被认为是靶基因(低氧诱导的pH无关)的基因，必须在四个细胞系中的三个中存在基因。结论。其结果是在由27个基因组成的细胞系中形成了一个不受pH影响的强健的缺氧谱。这项研究展示了一种利用微阵列识别缺氧标志物的方法，其中考虑了肿瘤微环境中的其他因素。
Background. Genes upregulated by low oxygen have been suggested as endogenous markers for tumor hypoxia. Yet, most of the genes investigated have shown inconsistent results, which have led to concerns about their ability to be true hypoxia makers. Previous studies have demonstrated that expression of hypoxia induced genes can be affected by extracellular pH (pH e). Methods. Five different human cell lines (SiHa, FaDu(DD), UTSCC5, UTSCC14 and UTSCC15) were exposed to different oxygen concentrations and pH (7.5 or 6.3), and gene expression analyzed with microarray (Affymetrix - Human Genome U133 Plus 2.0 Array). Results. An analysis of two of the cell lines using SAM identified 461 probesets that were able to separate the four groups "Normal oxygen, normal pH", "Low oxygen, normal pH", "Normal oxygen, low pH" and "Low oxygen, low pH". From here it was possible to identify a fraction of probesets induced at low oxygen independent of pH in these two cell lines, this fraction included HIG2, NDRG1, PAI1 and RORA. Further verification by qPCR highlighted the necessity of using more cell lines to obtain a robust gene expression profiles. To specifically select pH independent hypoxia regulated genes across more cell lines, data for FaDu(DD), UTSCC5, UTSCC14 and UTSCC15 were analyzed to identify genes that were induced by hypoxia in each cell line, where the induction was not affected by low pH, and where the gene was not significantly induced by low pH alone. Each cell line had 65-122 probesets meeting these criteria. For genes to be considered as target genes (hypoxia inducible pH independent), genes had to be present in three of four cell lines. Conclusion. The result is a robust hypoxia profile unaffected by pH across cell lines consisting of 27 genes. This study demonstrates a way to identify hypoxia markers by microarray, where other factors in the tumor microenvironment are taken into account.