胃癌通过激活Sonic Hedgehog通路诱导淋巴内皮细胞坏死性凋亡从而促进淋巴转移

At present, the EMT characteristic in tumor cells was important for the survival and infiltration of tumor cells in lymphatic vessel, but little is known about its mechanisms. We found out that gastric cancer cells (GCC) can synthetize and secrete Sonic Hedgehog（Shh）to maintain its EMT characteristic by autocrine action. Moreover, the co-culture of GCC and human lymphatic endothelial cells (HLEC) led to the the activation of RIP3/MLKL pathway and necroptosis in HLEC. The overexpression of Shh in HLEC resulted in the activation of MLKL. If the secreted Shh led to the overexpression of Shh in HLEC and necroptosis? We further found out that the silencing of Shh in GCC and neutralization of Shh in the co-culture system can both block down the activation of RIP3/MLKL and necroptosis in HLEC. We hypothesized that GCC secrete Shh and activate the Shh pathway in HLEC. The later led to the activation of RIP3/MLKL pathway and induced its necroptosis. We aimed to verify this hypothese in clinical sample, animal model and cellular level. It will give help to clarify the molecular mechanism of GCC lymphatic metastasis and provide a theoretical basis for individualized treatment of gastric cancer.

目前认为肿瘤细胞维持上皮间质化（EMT）是其在淋巴管中存活并侵出的关键，但对其侵出的机制知之甚少。我们的前期研究证实胃癌细胞（GCC）通过合成分泌Sonic Hedgehog（Shh）维持其EMT特性。最近发现GCC与人淋巴内皮细胞（HLEC）共培养可诱导后者RIP3/MLKL上调并发生坏死性凋亡；HLEC内Shh上调并介导了MLKL转录激活。是否GCC分泌的Shh导致HLEC内Shh上调并发生坏死性凋亡？进一步实验发现：中和共培养体系中游离Shh和沉默GCC Shh均可阻断HLEC的RIP3/MLKL上调及其坏死性凋亡。据此我们提出科学假设：GCC通过分泌Shh上调HLEC内Shh通路，后者上调HLEC内RIP3/MLKL并诱导其坏死性凋亡，从而破环淋巴管内皮屏障以利于侵出。本项目拟从临床样本、动物模型、细胞水平验证该假设，揭示胃癌淋巴转移机制，为胃癌靶向治疗提供新思路。

胃癌是我国威胁国民健康的重要疾病，胃癌的淋巴转移是存在于治疗中的一大难题，对胃癌淋巴转移的机制研究将有助于为临床工作提供新的思路。m6A修饰是广泛存在于细胞中的一种可逆性甲基化修饰，通过调控目标基因RNA的定位、翻译或稳定性对细胞的增殖转移等生物学行为产生影响。基于前期基础我们发现胃癌细胞分泌Shh蛋白可明显影响血管内皮细胞的增殖，诱导其凋亡；后续通过对m6A修饰相关蛋白研究，提出METTL3-PBX1-GCH1轴调控胃癌细胞四氢喋呤代谢影响其恶性进展，METTL14及ALKBH5对胃癌细胞恶行行为也具有明显调控作用，ALKBH5可明显促进胃癌细胞EMT通路激活；同时我们在胃癌中发现新的抑癌基因LCN2明显影响胃癌细胞转移能力。相关实验内容部分已经发表，相关结果对寻找胃癌新的治疗思路具有积极作用。目前项目仍缺少动物实验验证拮抗喋呤类物质生成作为治疗胃癌新靶向的有效性及安全性，后续将在次基础深入探索喋呤类物质对胃癌细胞代谢的影响，寻找四氢喋呤与细胞凋亡联系的关键。