微囊藻毒素-LR通过miR-221/IRX5调控EMT促进结直肠癌侵袭转移机制的研究

The data of our previous research (supported by NCFS NO. 21177062) showed that Microcystin-LR, the derivatives of cyanobacterial blooms，could enhance colorectal cancer invasion and metastasis. MiR-221 was a significantly up-regulated miRNA, which was screened by miRNA microarray using colorectal cancer cell line DLD-1 exposed to MC-LR. IRX5 was discovered as its target gene by bio-informatics analysis and luciferase reporter gene assay. The previous results and the related paper provided the evidence that miR-221 might play an import role in MC-LR induced EMT by its target gene IRX5, resulting in colorectal cancer invasion and metastasis. In this study, we will further investigate the roles and mechanisms of miR-221/ IRX5 on MC-LR-induced EMT, invasion and metastasis by not only using gene over-expression but also gene slience technology etc. Based on this, we will use clinic colorectal cancer to analysis the expression of miR-221 and IRX5. Our research will not only contribute to clarify the molecular mechanisms of MC-LR on colorectal cancer promotion, but also provide new scientific basis to prevent and cure cancer caused by environmental factors.

我们的前期研究（国家自然科学基金资助，No.21177062)表明，蓝藻水华衍生物微囊藻毒素-LR（MC-LR）可以促进结直肠癌侵袭转移。为深入探讨其机制，我们应用miRNA芯片分析了结直肠癌细胞DLD-1经MC-LR暴露后的miRNA变化，其中miR-221差异明显且与结直肠癌侵袭转移相关。生物信息学分析及荧光素酶报告基因检测发现，IRX5是miR-221的靶基因。前期结果和相关文献提示，MC-LR可能通过miR-221调控IRX5，促进EMT，进而促进结直肠癌侵袭转移。本研究拟以细胞及裸鼠为研究对象，利用基因过表达、基因沉默等技术研究miR-221及IRX5在MC-LR致结直肠癌EMT及侵袭转移中的作用与机制；在此基础上，利用临床结直肠癌样本分析miR-221、IRX5的表达。本研究不仅有助于阐明MC-LR促进结直肠癌的分子机制，还可为早期诊断和预防由环境因子导致的的癌症提供依据。

我们在前期研究中采用蓝藻水华衍生物微囊藻毒素-LR（Microcystin-LR, MC-LR）处理结直肠癌细胞DLD-1细胞，经miRNA 芯片检测，筛选出差异明显的miRNA，即hsa-miR-221；生物信息学分析和荧光素酶报告基因检测进一步筛选得到miR-221的候选靶基因IRX5。本研究拟探讨miR-221及IRX5在MC-LR致结直肠癌EMT及侵袭转移中的作用及其分子机制。我们发现，MC-LR可以通过PI3-K/AKT和SMAD2促进结直肠癌细胞发生EMT，进而导致肿瘤细胞侵袭、迁移能力增强；且这种促进结直肠癌细胞迁移能力增强是MC-LR通过上调肿瘤细胞中 miR-221实现的。但是慢病毒转染获得的IRX5稳定高表达的结直肠癌细胞系DLD-1表现出了明显的促进肿瘤转移的作用，与预期研究结果不一致，且IRX5促进结直肠癌转移作用与其调控RHOA信号通路显著相关。此外，IRX5还可以促进基因组不稳定性。因此，我们的实验调整为研究了MC-LR促进肿瘤转移是否通过调控miR-221的另外一个靶基因，即PTEN基因实现。我们发现，MC-LR可以通过miR-221/PTEN激活STAT3促进β-catenin入核，进而促进结直肠癌细胞迁移。本研究不仅有助于阐明MC-LR促进结直肠癌的分子机制，还可为早期诊断和预防由环境因子导致的的癌症提供依据。