MicroRNA-200b is downregulated in colon cancer budding cells

Background
The microRNA-200 (miR-200) family acts as a major suppressor of epithelial-mesenchymal transition (EMT). Impaired miR-200 expression may lead to EMT initiation and eventually cancer dissemination. The presence of tumor budding cells (TBC) is associated with metastasis and poor prognosis, and molecular similarities to EMT indicate that these cells may reflect ongoing EMT. The aim of this study was to investigate the expression of miR-200b in budding cells of colon cancer and the relationship with the EMT-markers E-cadherin, β-catenin and laminin-5γ2.
Material & methods
MiR-200b was investigated by in situ hybridization in 58 cases of stage II (n = 36) and III colon (n = 22) cancers with tumor budding. Expression of E-cadherin, β-catenin and laminin-5γ2 was examined by immunohistochemistry. A multiplex fluorescence assay combining miR-200b with cytokeratin and laminin-5γ2 was employed on a subset of 16 samples.
Results
MiR-200b was downregulated in the TBC at the invasive front of 41 out of 58 (71%) cases. The decline was present in both mismatch satellite stable and instable adenocarcinomas. The majority of cases also showed loss of membranous E-cadherin and increased nuclear β-catenin in the TBC, while laminin-5γ2 expression was upregulated at the invasive front and in the tumor buds of approximately half the adenocarcinomas. However, the miR-200b decline was not statistically associated with the expression of any of the EMT-markers. The miR-200b decline was also documented by multiplex fluorescence. Fourteen out of fifteen cases showed a decrease in miR-200b expression in the majority of the TBC, but no obvious relationship between miR-200b and laminin-5γ2 expression was observed. Conclusion: The findings support the assumption of a miR-200b related downregulation in colon cancer budding cells. Whether miR-200b expression may be of clinical significance awaits further studies.

【背景】微小RNA-200（microRNA-200, miR-200）家族是上皮间质转化（epithelial-mesenchymal transition, EMT）的主要抑制因子。miR-200表达异常可诱发EMT进程，最终促进肿瘤播散。肿瘤出芽细胞（tumor budding cells, TBC）的存在与肿瘤转移及不良预后密切相关，且其分子特征与EMT高度相似，提示此类细胞正处于EMT进程中。本研究旨在探讨miR-200b在结肠癌肿瘤出芽细胞中的表达情况，及其与EMT标志物E-钙粘蛋白、β-连环蛋白及层粘连蛋白-5γ2的相关性。
【材料与方法】选取58例伴肿瘤出芽的结肠癌患者标本，其中Ⅱ期36例、Ⅲ期22例，采用原位杂交技术检测miR-200b的表达；采用免疫组织化学法检测E-钙粘蛋白、β-连环蛋白及层粘连蛋白-5γ2的表达。另外选取16例标本开展联合检测，采用多重荧光检测技术同时标记miR-200b、细胞角蛋白及层粘连蛋白-5γ2。
【结果】58例标本中，41例（71%）的侵袭前沿肿瘤出芽细胞内miR-200b表达下调，且该现象在错配卫星稳定型及错配卫星不稳定型腺癌中均存在。多数标本的肿瘤出芽细胞还可见膜型E-钙粘蛋白缺失及核内β-连环蛋白表达升高；约半数腺癌的侵袭前沿及肿瘤出芽区域可见层粘连蛋白-5γ2表达上调。但miR-200b表达下调与上述任一EMT标志物的表达均无统计学相关性。多重荧光检测进一步验证了miR-200b的下调现象：15例标本中14例的多数肿瘤出芽细胞内miR-200b表达降低，但未观察到miR-200b与层粘连蛋白-5γ2表达存在明显关联。
【结论】本研究结果支持“结肠癌肿瘤出芽细胞中存在miR-200b相关表达下调”这一假说。miR-200b的表达是否具有临床意义，尚需后续进一步研究阐明。