miR-100通过胞内及胞外途径调控SWI/SNF复合物影响大肠癌放射敏感性的机制研究

The prognosis of colon cancer induced by radioresistance remains a concern. Radiosensitivity of colon cancer is closely related to abnormal expression of miRNAs, EMT pathway genes and tumor microenvironment. MiRNA influence the radiosensitivity through regulating the EMT pathway,and the content of miRNA may changes because of exosome-mediated pathways in tumor microenvironment. We have found that miR-100 was responsive to X-ray irradiation and conferred increased radiosensitivity to colon cancer cells. Bioinformatic tools have predicted that miR-100 can regulate EMT pathway SWI/SNF.There is no similar report about miR-100 regulating the radiosensitivity of colon cancer and exosome-mediated pathways. SWI/SNF pathway in EMT is a new functional model.Therefore, we postulated that miR-100 can regulate the radiosensitivity of colon cancer cells via targeting EMT through cytoplasmic and exosome-mediated pathways. This project aims to illustrate the mechanism how miR-100 regulate radiosensitivity of colon cancer cells via targeting EMT. We will use various molecular biology methods to 1) investigate the target genes of miR-100; 2) explore the effects of miR-100 on radiosensitivity; 3) explore the effect of CAF miR-100 expression on co-cultured colon cancer. Moreover, we discuss the function of miR-100 on the radiosensitivity of colon cancer via exosome-mediated pathways. This study aims to illustrate the mechanism of miR-100 influencing the radiosensitivity of colon cancer binding cytoplasmic with exosome-mediated pathways.

放疗抵抗对大肠癌预后的影响是临床面临的大问题。大肠癌细胞放射敏感性与miRNA的异常表达、上皮间质转化（ EMT）及肿瘤相关成纤维细胞（CAF）等微环境因素密切相关。miRNA可通过调控EMT影响肿瘤细胞放射敏感性，而肿瘤微环境中外泌体的介导则可改变肿瘤细胞内miRNA水平。本课题预实验通过二代测序和克隆试验发现miR-100参与了大肠癌细胞放射抵抗，生物信息学表明miR-100可调控EMT通路SWI/SNF复合物家族的多个关键分子。miR-100对大肠癌放射敏感性的调控及外泌体介导下的功能目前未见报道。本项目拟在细胞和动物水平上，研究miR-100调控EMT通路靶基因从而影响大肠癌放射敏感性的机制。同时研究CAF通过外泌体中miR-100调控大肠癌放射敏感性的作用模式。本研究将揭示miR-100通过细胞内和细胞外途径影响大肠癌放射敏感性的机制，有望阐明大肠癌放射敏感性新的途径。

大肠癌细胞放射敏感性与miRNA的异常表达，上皮间质转化（EMT）和肿瘤相关成纤维细胞（CAF）等微环境因素密切相关。 miRNA可通过调控EMT影响肿瘤细胞放射敏感性，而肿瘤微环境中外泌体的介导可可改变肿瘤细胞内miRNA水平。本课题预实验通过二代测序和克隆试验发现miR-100参与了大肠癌细胞放射抵抗，生物信息学表明miR-100可调控EMT通路SWI / SNF复合物家族的多个关键分子。我们研究发现大肠癌组织中miR-100明显低于与其匹配的非癌性结肠组织。miR-100的上调可通过抑制细胞的增殖，克隆形成，促进凋亡以及DNA双链断裂和抑制EMT的发生来增加CCL-244的辐射敏感性。在大肠癌CCL-244细胞中上调miR-100可以抑制抑制SWI / SNF复合物的表达，反之则促进其表达，这与在CCL-244的辐射调节结果一致。利用组织芯片进行免疫组化检测SWI/SNF复合物的表达发现SMARCA4，SMARCA5在肿瘤组织中高表达，在癌旁正常组织中低表达或无表达。在动物水平，上调miR-100后进行照光发现与对照组相比其肿瘤生长受到抑制。另外，我们研究发现MiR-31对CAF的表型，增殖，周期以及凋亡均无影响，但是miR-31上调可以明显抑制CAF的自噬，进而影响与miR-31调节后的CAF共培养的大肠癌细胞的辐射敏感性。这与我们的课题组之前研究的结果：CAF自噬抑制后可以明显增加大肠癌的辐射敏感性相一致。综上所述，miR-100可以明显调节大肠癌的辐射敏感性，这一作用可能通过SWI / SNF通路实现的。另外,miR-31调节CAF自噬状态并影响大肠癌的生物学行为。