幽门螺杆菌调节环状RNAhsa_circ_0002669表达促进胃上皮细胞恶性转化的分子机制

Infection with helicobacter pylori (H. pylori) is regarded as the strongest risk factor for gastric carcinogenesis and progression. CircRNA is considered as the new research hotspots in the field of RNA. Our previous study results showed that the level of hsa_circ_0002669 was down-regulated significantly in human gastric cancer tissues compared with the corresponding noncancerous tissues by RNA sequencing analysis and qRT-PCR validation. Furthermore, we found that the level of hsa_circ_0002669 can be significantly inhibited with helicobacter pylori infection in gastric epithelial cells. Overexpression of hsa_circ_0002669 can inhibited cell proliferation and migration significantly. But the molecular mechanism has not yet been clarified. Based on the recent research progress and our previous studies, the aim of this project is to explore the molecular mechanism of gastric mucosa tissues malignant transformation induced helicobacter pylori by regulating hsa_circ_0002669 from molecules and cells level, experimental animal level and the clinical gastric tissue sample level. The main contents of this project are as follows： (1) To investigate whether helicobacter pylori can up-regulate RNA editing enzymes ADAR1 expression through NF-κB andβ-catenin signal pathway. To make clear the molecular mechanism of helicobacter pylori inhibiting hsa_circ_0002669 expression from RNA editing angle mediated by ADAR1；(2) To make clear the molecular mechanisms of hsa_circ_0002669 inhibiting the malignant transformation of gastric epithelial cells. To investigate whether hsa_circ_0002669 can act as miRNA molecule "sponge" and identify the miRNA that can bind with hsa_circ_0002669 and explore the downstream target molecules of the miRNA. To elucidate the mechanism of helicobacter pylori promoting the malignant transformation by regulating hsa_circ_0002669 and its downstream target molecules network.(3) To verify that helicobacter pylori promotes the malignant transformation of gastric mucosa tissues by regulating hsa_circ_0002669 and its downstream target molecules network from the animal level and human pathological tissue levels. This study aims to provide the theory evidence for the hypothesis that hsa_circ_0002669 can be used as a key point and therapeutic target in the early intervention of gastric cancer .

幽门螺杆菌感染是导致胃上皮细胞癌变最强的危险因素，环状RNA是目前RNA领域的最新研究热点，申请者首次发现环状RNAhsa_circ_0002669在胃癌组织中表达显著降低并发挥重要的生物学功能，幽门螺杆菌感染可显著抑制其表达。本课题拟在此基础上：①探讨幽门螺杆菌通过NF-κB及β-catenin依赖的途径调节RNA编辑酶ADAR1表达，并从ADAR1介导的RNA编辑视角解析幽门螺杆菌抑制该环状RNA表达的分子机制；②确定该环状RNA作为miRNA“海绵”所吸附的miRNA并剖析miRNA所靶向的下游分子，明确该环状RNA抑制胃粘膜细胞恶性转化的分子机制，阐明幽门螺杆菌通过调控该环状RNA及其下游分子网络促进细胞恶性转化的机理。③从动物及临床组织标本中验证该环状RNA及其分子网络在幽门螺杆菌感染促进胃粘膜细胞恶性转化中的作用。为基于circ_0002669为靶点的胃癌早期干预提供理论依据。

幽门螺杆菌感染是导致胃上皮细胞癌变最强的危险因素，环状RNA在肿瘤的发生发展过程中发挥着关键性的调控作用。本项目我们探讨了幽门螺杆菌对环状RNAhsa_circ_0002669及hsa_circ_0004872表达的调控及这两种环状RNA在胃癌中的表达、调控及作用机制。我们发现幽门螺杆菌感染胃癌细胞后，可通过β-catenin依赖的信号途径降低hsa_circ_0002669的表达，而对hsa_circ_0004872的表达没有显著影响。临床组织标本检测发现：这两种环状RNA在胃癌中的表达水平均显著降低，而且二者的表达水平与肿瘤大小、局部淋巴结转移等密切相关。将二者的表达载体分别转染胃癌细胞后，细胞的增殖能力及侵袭转移能力均显著降低，反之，利用特异性的siRNA将二者干扰后细胞的增殖及侵袭转移能力则明显增强。利用裸鼠体内实验我们也证实了这两种环状RNA的抑癌功能。进一步的分子机制研究发现hsa_circ_0002669可作为miR-223-3p的分子海绵，并通过miR-223-3p/Arid1a轴调控胃癌的发生发展。而hsa_circ_0004872可作为miR-224的分子海绵并进一步上调miR-224所调控的下游靶分子p21及Smad4的表达，而且RNA编辑酶ADAR1可调控hsa_circ_0004872的表达，并通过hsa_circ_0004872/miR-224轴抑制Smad4的表达水平，而Smad4可进一步结合在ADAR1的启动子上来抑制其表达，因此，hsa_circ_0004872可通过miR-224/Smad4/ADAR1/hsa_circ_0004872这样一个连续的负调控环路来抑制胃癌进展。本研究丰富了对幽门螺杆菌致病机制的认识，并为以hsa_circ_0002669及hsa_circ_0004872为靶点的胃癌早期诊断及治疗提供了理论依据。