Rechreche H, Montalto G, Mallo G V, Vasseur S, Marasa L, Soubeyran P, Dagorn J C, Iovanna J L
U.315 INSERM, Marseille, France.
Int J Cancer. 1999 May 31;81(5):688-94. doi: 10.1002/(sici)1097-0215(19990531)81:5<688::aid-ijc3>3.0.co;2-r.
We have established the phenotype of a colorectal tumor by partial sequencing of 2166 transcripts that were eventually arrayed on high-density filters. These filters were used for differential screening with mRNAs of colorectal cancer and normal adjacent mucosa to characterize genes whose expression is altered in colorectal carcinoma. Three genes encoding related proteins, PAP, reg Ialpha and reg Ibeta, were over-expressed in cancer. Northern-blot analysis confirmed that their expression was very low in normal colonic epithelial cells, but elevated in 75% of tumors. Western blotting with specific antibodies to pap and reg Ialpha revealed in tumors a single band of the expected size ( 15-16 kDa), demonstrating synthesis of the proteins. Pap was localized by immunohistochemistry to the cytoplasm of epithelial cells. In cancerous tissue, many cells showed a strong staining signal, but the proportion of stained cells was variable among patients. In normal mucosa, staining was light and restricted to a few cells scattered in the epithelium. Similar results were obtained with antibodies against reg Ialpha. No significant relationship was found between concentrations of pap, reg Ialpha or reg Ibeta and clinical outcome. We looked at potential effectors of pap/reg gene over-expression by testing, in 2 adenocarcinoma cell lines, the efficacy of the pap promoter at driving a reporter gene; strong induction was observed upon exposure to IFNgamma and IL-6. By analogy with observations in hepatocellular carcinoma, our results suggest that prevention of PAP/reg expression in normal colon cells by silencing their gene promoters is relieved during colon carcinogenesis, allowing their up-regulation by mediators such as cytokines.
我们通过对2166个转录本进行部分测序，确定了结肠直肠肿瘤的表型，这些转录本最终被排列在高密度滤膜上。这些滤膜用于与结肠直肠癌和正常相邻黏膜的mRNA进行差异筛选，以鉴定在结肠直肠癌中表达发生改变的基因。三个编码相关蛋白的基因，即PAP、reg Iα和reg Iβ，在癌症中过度表达。Northern印迹分析证实，它们在正常结肠上皮细胞中的表达非常低，但在75%的肿瘤中升高。用针对pap和reg Iα的特异性抗体进行蛋白质印迹分析，在肿瘤中显示出一条预期大小（15 - 16 kDa）的单带，证明了这些蛋白质的合成。通过免疫组织化学将Pap定位到上皮细胞的细胞质中。在癌组织中，许多细胞显示出强烈的染色信号，但不同患者之间染色细胞的比例各不相同。在正常黏膜中，染色较浅，仅限于上皮中散在的少数细胞。用针对reg Iα的抗体也得到了类似的结果。未发现pap、reg Iα或reg Iβ的浓度与临床结果之间存在显著关系。我们通过在2种腺癌细胞系中测试pap启动子驱动报告基因的效力，来研究pap/reg基因过度表达的潜在效应物；在暴露于IFNγ和IL - 6时观察到强烈的诱导作用。与肝细胞癌中的观察结果类似，我们的结果表明，在结肠癌发生过程中，通过沉默其基因启动子来预防正常结肠细胞中PAP/reg表达的抑制作用被解除，从而使其能够被细胞因子等介质上调。
