de Bruin Elza C, van de Pas Simone, Lips Esther H, van Eijk Ronald, van der Zee Minke M C, Lombaerts Marcel, van Wezel Tom, Marijnen Corrie A M, van Krieken J Han J M, Medema Jan Paul, van de Velde Cornelis J H, Eilers Paul H C, Peltenburg Lucy T C
Department of Clinical Oncology, Leiden University Medical Center, Albinusdreef 2, 2333 ZA Leiden, The Netherlands.
BMC Genomics. 2005 Oct 14;6:142. doi: 10.1186/1471-2164-6-142.
The molecular determinants of carcinogenesis, tumor progression and patient prognosis can be deduced from simultaneous comparison of thousands of genes by microarray analysis. However, the presence of stroma cells in surgically excised carcinoma tissues might obscure the tumor cell-specific gene expression profiles of these samples. To circumvent this complication, laser microdissection can be performed to separate tumor epithelium from the surrounding stroma and healthy tissue. In this report, we compared RNAs isolated from macrodissected, of which only surrounding healthy tissue had been removed, and microdissected rectal carcinoma samples by microarray analysis in order to determine the most reliable approach to detect the expression of tumor cell-derived genes by microarray analysis.
As microdissection yielded low tissue and RNA quantities, extra rounds of mRNA amplification were necessary to obtain sufficient RNA for microarray experiments. These second rounds of amplification influenced the gene expression profiles. Moreover, the presence of stroma cells in macrodissected samples had a minor contribution to the tumor cell gene expression profiles, which can be explained by the observation that more RNA is extracted from tumor epithelial cells than from stroma.
These data demonstrate that the more convenient procedure of macrodissection can be adequately used and yields reliable data regarding the identification of tumor cell-specific gene expression profiles.
致癌作用、肿瘤进展和患者预后的分子决定因素可通过微阵列分析同时比较数千个基因来推导。然而，手术切除的癌组织中存在基质细胞可能会掩盖这些样本中肿瘤细胞特异性基因表达谱。为了规避这一复杂情况，可进行激光显微切割以将肿瘤上皮与周围基质和健康组织分离。在本报告中，我们通过微阵列分析比较了从仅去除周围健康组织的大块解剖样本以及显微切割的直肠癌样本中分离的RNA，以确定通过微阵列分析检测肿瘤细胞衍生基因表达的最可靠方法。
由于显微切割产生的组织和RNA量较低，需要额外轮次的mRNA扩增才能获得足够的RNA用于微阵列实验。这些第二轮扩增影响了基因表达谱。此外，大块解剖样本中基质细胞的存在对肿瘤细胞基因表达谱的影响较小，这可以通过观察到从肿瘤上皮细胞中提取的RNA比从基质中提取的更多来解释。
这些数据表明，更便捷的大块解剖程序可以得到充分利用，并能产生关于识别肿瘤细胞特异性基因表达谱的可靠数据。
