Analysis of the invasive capacity at the invasive front of colon cancer, using crypt isolation technique and microarray

为了分析癌细胞的侵袭能力，我们筛选了在侵袭前沿表达特异性变化的基因。我们从20个手术切除的肿瘤样本中收集了侵袭性前部和肿瘤中心的癌细胞。虽然我们一开始尝试了隐窝分离技术，但很难从每个部位正确分离和收获癌细胞。因此，我们通过激光显微解剖技术采集癌细胞，进行微阵列分析，并确定了在侵袭前沿表达特异性变化的基因。通过Student-t检验，我们发现5个基因在侵入前信号上调，1个基因信号下调。聚类分析显示，24个基因上调，29个基因下调。此外，最近的一些报道显示，侵袭前存在低分化的肿瘤与预后不良有关。在11例具有该组织学特征的病例中，我们可以通过Student-t检验阐明4个上调基因和6个下调基因。通过聚类分析，共鉴定出13个上调基因和18个下调基因。我们的研究结果表明，许多与蛋白水解或细胞外基质无关的分子可能与癌细胞侵袭或分化不良有关。我们现在正在用RT-PCR确认所拾取基因的表达。

To analyze the invasive capacity of cancer cells, we screened the genes whose expression specifically changes at the invasive front. We harvested the cancer cells at the invasive front and the center of the tumor from 20 surgically resected samples. Though we attempted the crypt isolation technique at first, it was very difficult to isolate and harvest cancer cells from each part correctly. So we harvested cancer cells by laser micro-dissection technique, conducted microarray analysis, and identified the genes whose expression changed specifically at the invasive front. By Student-t test, we picked up five genes whose signals were up-regulated at the invasive front, and one gene whose signal was down-regulated. By cluster analysis, twenty-four genes were up-regulated, and twenty-nine genes were down-regulated. Moreover, some recent reports revealed that the presence of poorly differentiated clusters at the invasive front was correlated with poor prognosis.When examined the eleven cases with this histological feature, we could elucidate four up-regulated and six down-regulated genes by Student-t test. By cluster analysis, thirteen up-regulated and eighteen down-regulated genes were identified.Our results suggested that many molecules not related with proteolysis or extra-cellular matrix might correlate with cancer cell invasion or poor differentiation. We are now confirming expression of the picked-up genes by RT-PCR.