Department of Biochemistry and Molecular Biology, University of North Dakota, Grand Forks, North Dakota; The State Key Laboratory for Biotherapy, West China Hospital, Sichuan University, Chengdu, China.
Gastroenterology. 2013 Oct;145(4):853-64.e9. doi: 10.1053/j.gastro.2013.06.008. Epub 2013 Jun 13.
BACKGROUND & AIMS: Little is known about functions of microRNA (miR) passenger strands (miR*) or their roles in tumor development or progression. We screened for miRs and miR* with levels that were altered in metastatic colorectal cancer (CRC) cells and human tumor samples and investigated their targets and effects on cell function and tumor progression in mice.
We performed array-based profile analysis to identify miRs with levels that were increased more than 2-fold in metastatic (SW620) CRC cells compared with nonmetastatic (SW480) cells. Quantitative polymerase chain reaction and in situ hybridization analyses were used to measure miRNA levels in CRC cell lines and human tumor samples. We used miRNA duplex mimics or inhibitors to increase and decrease levels of miRNA in CRC cells and assessed their activities and ability to form metastatic xenograft tumors in nude mice.
Levels of miR-221* and miR-224 were reduced in metastatic compared with nonmetastatic CRC cells; levels in human tumor samples correlated inversely with tumor stage and metastasis to lymph nodes as well as patient survival times. SW480 cells transfected with miR-221* or miR-224 inhibitors had increased motility in vitro compared with SW480 control cells and formed larger, more metastatic tumors when injected into mice. SW620 cells transfected with miR-221* or miR-224 mimics had reduced migration and motility in vitro and formed smaller tumors with fewer metastases in mice compared with control SW620 cells. We identified the 3' untranslated region of MBD2 messenger RNA as a target of miR-221* and miR-224. MBD2 silences the gene encoding maspin, a suppressor of metastasis. In CRC cells, we found that miR-221* and miR-224 increase the expression of maspin through MBD2 down-regulation.
In metastatic CRC cells, reduced levels of miR-221* and miR-224 increase levels of MBD2, thereby decreasing expression of the metastasis suppressor maspin. Increased activities of miR-221* and miR-224 reduce growth and metastasis of CRC xenograft tumors in mice; these miRs might be developed as therapeutic reagents or biomarkers of CRC progression.
关于微 RNA (miR) 过客链 (miR*) 的功能或它们在肿瘤发生和发展中的作用知之甚少。我们筛选了在转移性结直肠癌 (CRC) 细胞和人类肿瘤样本中水平改变的 miR 和 miR*，并研究了它们在小鼠中的靶标和对细胞功能及肿瘤进展的影响。
我们进行了基于阵列的谱分析，以鉴定在转移性 (SW620) CRC 细胞中水平增加超过 2 倍的 miR，与非转移性 (SW480) 细胞相比。定量聚合酶链反应和原位杂交分析用于测量 CRC 细胞系和人类肿瘤样本中的 miRNA 水平。我们使用 miRNA 双链体模拟物或抑制剂来增加和减少 CRC 细胞中的 miRNA 水平，并评估它们在裸鼠中的活性和形成转移性异种移植肿瘤的能力。
与非转移性 CRC 细胞相比，转移性 CRC 细胞中 miR-221* 和 miR-224 的水平降低；人类肿瘤样本中的水平与肿瘤分期和淋巴结转移以及患者生存时间呈负相关。与 SW480 对照细胞相比，转染 miR-221* 或 miR-224 抑制剂的 SW480 细胞在体外具有更高的迁移率，并在注射到小鼠中时形成更大、更具转移性的肿瘤。与对照 SW620 细胞相比，转染 miR-221* 或 miR-224 模拟物的 SW620 细胞在体外迁移和运动能力降低，在小鼠中形成的肿瘤较小，转移较少。我们鉴定了 MBD2 信使 RNA 的 3'非翻译区作为 miR-221* 和 miR-224 的靶标。MBD2 沉默了 maspin 的基因，maspin 是一种抑制转移的基因。在 CRC 细胞中，我们发现 miR-221* 和 miR-224 通过下调 MBD2 增加 maspin 的表达。
在转移性 CRC 细胞中，miR-221* 和 miR-224 的水平降低增加了 MBD2 的水平，从而降低了转移抑制因子 maspin 的表达。miR-221* 和 miR-224 的活性增加减少了 CRC 异种移植肿瘤在小鼠中的生长和转移；这些 miR 可能被开发为 CRC 进展的治疗试剂或生物标志物。
