环状RNA靶向调控miR-3663-3p/ID3信号轴在AFB1致细胞恶性转化中的作用机制研究

The development of lung cancer is closely identified with air pollution. Airborne AFB1 has been classified as a potent carcinogen which can be bioactivated by the cytochrome P450 2A13. However, the mechanism of this action is unclear. In our previous research, we constructed an immortalized human bronchial epithelial cell (BEAS-2B) stably expressing CYP2A13 (B-2A13 cells). As a sensitive cell model, it accelerated the cell malignant transformation processed by metabolic activation of AFB1 to produce ultimate carcinogens. Importantly, we can observe the dynamic process of the whole cell malignant transformation in real time. Our previous study showed that miR-3663-3p/ID3 regulatory axis plays an important role in AFB1-induced cell malignant transformation, which screened by mRNA chip and miRNA chip. At present, more and more researchers were devoted to the study on the role of circRNA-miRNA-mRNA axis in bioscience area. However, the role of circRNA in the cell malignant transformation needs to be emphasized. In this study, we identified up-regulated miRNA expression between P50 B-2A13 cells and P50 B-Vector cells by microarray analysis, screening those circRNAs that are associated with malignant transformation of cells and building the cricRNA/miR-3663-3p targeting relationship library. Then, the effect and mechanism of circRNA/hsa-miR-3663-3p/ID3 axis on cell function are detected. In vivo function verification, we will use the malignant transformation cells of different generations and tumor xenograft transplantation into nude mice and zebrafish to clarify the role of the regulatory axis in the whole process of malignant transformation of cells. The present study will be helpful to strengthen the understanding of AFB1 in relating to lung cancer, and provide biomarkers and strategy in prediction, therapy and prevention of AFB1-induced respiratory system cancer.

肺癌的发生与环境空气污染密切相关，AFB1是重要的空气污染物，CYP2A13酶可以代谢活化AFB1产生致癌作用，具体作用机制尚不明确。我们的前期研究构建代谢酶高表达细胞系（B-2A13）作为敏感细胞模型，可加快AFB1致细胞恶转进程并易于观察全过程。通过mRNA芯片、miRNA芯片和细胞功能验证出miR-3663-3p/ID3轴在AFB1致细胞恶转中发挥重要作用。当前circRNA-miRNA-mRNA调控轴是研究热点，而circRNA在环境化学物所致细胞恶转中的作用研究较少。本研究拟利用已有的circRNA芯片构建cricRNA/miR-3663-3p靶向关系库，筛选出候选RNA调控轴再通过细胞功能和移植瘤模型验证其在细胞恶转过程中的作用，研究结果将有助于完善AFB1致呼吸系统肿瘤发生的机制，为探寻AFB1致呼吸系统肿瘤标志物提供新的研究策略。

肺癌的发生与环境空气污染密切相关，黄曲霉毒素AFB1是重要的空气污染物，CYP2A13酶可以代谢活化AFB1产生致癌作用，具体作用机制尚不明确。我们的前期研究构建代谢酶高表达细胞系（B-2A13）作为敏感细胞模型，可加快AFB1致细胞恶转进程并易于观察全过程。环状RNA(circRNA)作为一种新型非编码RNA，研究的最多的是“miRNA分子海绵”功能，可与mRNA竞争性结合miRNA进而影响基因的转录调控水平，形成以circRNA-miRNA-mRNA为轴的调控网络，而circRNA在环境化学物所致细胞恶转中的作用研究较少。本研究通过mRNA芯片、miRNA芯片和细胞功能验证出miR-3663-3p/ID3轴在AFB1致细胞恶转中发挥重要作用。本研究利用已有的circRNA芯片构建cricRNA/miR-3663-3p靶向关系库，筛选出候选RNA调控轴再通过细胞功能和移植瘤模型验证其在细胞恶转过程中的作用，研究结果最终阐明has_circ_0041714/hsa-miR-3663-3p/ID3调控轴调控轴在整个AFB1致人支气管上皮细胞的恶转过程中发挥的作用。.在该研究基础之上，外泌体miRNAs是一类新的细胞信使，在细胞之间穿梭并介导细胞间的物质转运与信息交流，参与机体稳态和疾病的调节。因此，我们探索性地去验证本基金项目发现的hsa-miR-3663-3p在外泌体中表达情况，但结果表明不太适合作为生物标志物。鉴于外泌体miRNAs的稳定性和miRNA基因簇的对功能蛋白的协同调控作用，所以我们通过一系列分子生物学实验和人群肺腺癌血浆、组织样本验证了外泌体型miR-487a/miR-487b簇在AFB1诱导的BEAS-2B细胞恶性转化进程中的作用。所有这些结果将有助于完善AFB1致呼吸系统肿瘤发生的机制，为探寻AFB1致呼吸系统肿瘤标志物提供新的研究策略。