Department of Biochemistry, Genetics and Immunology, University of Vigo, Campus As Lagoas Marcosende s/n, 36310 Vigo, Spain.
Int J Mol Sci. 2013 Aug 19;14(8):16986-98. doi: 10.3390/ijms140816986.
In previous studies we described a decreased alpha-L-fucosidase activity in colorectal tumors, appearing as a prognostic factor of tumoral recurrence. The aim of this work was to extend the knowledge about tissue alpha-L-fucosidase in colorectal cancer by quantifying the expression of its encoding gene FUCA1 in tumors and healthy mucosa. FUCA1 mRNA levels were measured by RT-qPCR in paired tumor and normal mucosa tissues from 31 patients. For the accuracy of the RT-qPCR results, five candidate reference genes were validated in those samples. In addition, activity and expression of alpha-L-fucosidase in selected matched tumor and healthy mucosa samples were analyzed. According to geNorm and NormFinder algorithms, RPLP0 and HPRT1 were the best reference genes in colorectal tissues. These genes were used for normalization of FUCA1 expression levels. A significant decrease of more than 60% in normalized FUCA1 expression was detected in tumors compared to normal mucosa (p = 0.002). Moreover, a gradual decrease in FUCA1 expression was observed with progression of disease from earlier to advanced stages. These findings were confirmed by Western blot analysis of alpha-L-fucosidase expression. Our results demonstrated diminished FUCA1 mRNA levels in tumors, suggesting that expression of tissue alpha-L-fucosidase could be regulated at transcriptional level in colorectal cancer.
在之前的研究中，我们描述了结直肠肿瘤中α-L-岩藻糖苷酶活性降低，其似乎是肿瘤复发的预后因素。本研究旨在通过定量检测肿瘤和正常黏膜组织中其编码基因 FUCA1 的表达，进一步了解结直肠癌组织中的α-L-岩藻糖苷酶。从 31 名患者的配对肿瘤和正常黏膜组织中，通过 RT-qPCR 测定 FUCA1 mRNA 水平。为了确保 RT-qPCR 结果的准确性，在这些样本中验证了 5 个候选参考基因。此外，还分析了选定的匹配肿瘤和正常黏膜样本中α-L-岩藻糖苷酶的活性和表达。根据 geNorm 和 NormFinder 算法，RPLP0 和 HPRT1 是结直肠组织中最好的参考基因。这些基因用于 FUCA1 表达水平的归一化。与正常黏膜相比，肿瘤中归一化 FUCA1 表达显著降低超过 60%（p = 0.002）。此外，随着疾病从早期到晚期的进展，FUCA1 表达逐渐降低。这些发现通过α-L-岩藻糖苷酶表达的 Western blot 分析得到了证实。我们的结果表明肿瘤中 FUCA1 mRNA 水平降低，提示结直肠癌中组织α-L-岩藻糖苷酶的表达可能受转录水平调控。
