lncRNA-C21orF96/miR-875-5p/USF2调控轴介导胃癌淋巴管转移的机制

Gastric cancer is mainly through lymph node metastasis, and the mechanism is not fully understood. In our previous study, we have found lncRNA-C21orF96 is associated with lymph node metastasis and tube formation. C21orF96 inhibits the expression of miR-875-5p, however, miR-875-5p targets USF2 gene, C21orF96 is involved with the regulation of USF2 gene. We speculate that lncRNA-C21orF96 regulates the lymphatic metastasis of gastric cancer by regulating miR-875-5p/USF2 via "Molecular sponge" function. In this study, we demonstrate the expression of C21orF96, miR-875-5p and USF2 in cancer tissues, blood and metastatic node of patients with gastric pathology, and their relationship with metastases associated characteristics and prognosis; we will try to construct a malignant lymphangiogenesis model to verify the function of lncRNA-C21orF96 in human gastric cancer and the lymphangiogenesis of gastric cancer; we explore the mechanisms of lncRNA-C21orF96/miR-875-5p/USF2 axis in the lymphatic metastasis of gastric cancer via “molecular sponge ”function using cell and animal models. In general, we hope to clarify the new mechanisms of LncRNA-C21orF96 in lymph metastases by miR-875-5p/USF2 pathway, and provide new target for the therapeutic interventions of gastric cancer with metastatic lymph nodes.

胃癌主要经淋巴结转移，其机制尚未完全阐明。我们前期发现lncRNA-C21orF96与胃癌淋巴转移和管形成相关；C21orF96抑制 miR-875-5p 表达，而miR-875-5p 直接靶向 USF2基因，参与 USF2 的表达调控，我们推测C21orF96 可能以“分子海绵”作用调控miR-875-5p/USF2 介导胃癌淋巴管的转移。本课题拟阐明C21orF96、miR-875-5p和USF2在胃癌组织、血液和转移灶中表达与胃癌病理特征和预后的关系；构建人高侵袭性胃癌淋巴管模型以明确C21orF96在胃癌和胃癌淋巴管形成的作用；以细胞和动物为模型探索C21orF96/miR-875-5p/USF2轴以“分子海绵”作用介导胃癌淋巴管转移的机制。通过上述研究明确lncRNA-C21orF96/miR-875-5p/USF2通路介导胃癌淋巴管转移的新机制，为胃癌治疗淋巴结转移提供新靶点。

目的：胃癌主要经淋巴结转移，其机制尚未完全阐明。我们前期发现lncRNA-C21orF96与胃癌淋巴转移和管形成相关；C21orF96抑制miR-875-5p表达，而miR-875-5p直接靶向USF2基因，参与USF2的表达调控，我们推测C21orF96可能以“分子海绵”作用调控miR-875-5p/USF2介导胃癌淋巴管的转移。本课题拟：1)阐明C21orF96、miR-875-5p和USF2在胃癌组织、血液和转移灶中表达与胃癌病理特征和预后的关系；2)构建人高侵袭性胃癌淋巴管模型以明确C21orF96在胃癌和胃癌淋巴管形成的作用；3)以细胞和动物为模型探索C21orF96/miR-875-5p/USF2轴以“分子海绵”作用介导胃癌淋巴管转移的机制，为胃癌治疗淋巴结转移提供新靶点。.方法：①用RT-PCR方法检测C21orF96、miR-875-5p在胃正常黏膜上皮细胞、胃癌细胞、阳性淋巴结组织及胃癌组织的表达情况。②用RT-PCR扩增C21orF96 cDNA全长片段，经过酶切测序验证后装入表达载体，再次酶切测序验证，用同样的方法获得干扰载体。③载体转染胃癌细胞，转染观察C21orF96对miR-875-5p和USF2的影响。④载体转染胃癌细胞，转染观察C21orF96对胃癌细胞侵袭、迁移和血管形成的影响。.结果：①RT-PCR结果显示C21orF96基因在胃正常粘膜上皮呈现低或不表达，而在阳性淋巴结组织、胃癌组织和胃癌细胞株中高表达，C21orF96与miR-875-5p呈负相关。②成功构建C21orF96的表达载体并转染胃癌细胞株，筛选出稳定表达的细胞，RT-PCR和Western blot证实实验组C21orF96表达上调。用同样的方法进行干扰载体转染。③C21orF96的表达上调，引起miR-875-5p表达显著下调而usf2基因表达升高，反之亦然。④C21orF96的过表达促进胃癌细胞的侵袭、迁移、小管形成。⑤C21orF96以“分子海绵”作用调控miR-875-5p/USF2介导胃癌淋巴管的转移。⑥C21orF96/miR-875-5p/USF2轴与胃癌患者淋巴结转移和TNM分期密切相关而与性别、年龄。肿瘤大小、细胞分化、癌细胞浸润深度、肿瘤部位和远处转移无关.结论：C21orF96/miR-875-5p/USF2轴与胃癌患者转移和预后密切相关。