幽门螺杆菌感染促进肿瘤相关成纤维细胞与胃癌细胞的互作及机制研究

Inflammatory microenvironment induced by Helicobacter pylori (H.pylori) plays an important role in the development and progression of gastric cancer. Cancer-associated fibroblasts (CAFs) constitute the major cell population in tumor microenvironment. We found that H.pylori can actively adhere to CAFs, and CAFs can promote the clonal growth of gastric cancer cells. The gerbil model of gastric cancer induced by H.pylori is accompanied by a lot of lymphocyte infiltration. Therefore, we speculate that the inflammatory response to H.pylori can directly and indirectly activate fibroblasts to CAFs. The activated CAFs secret many kinds of factors to establish inflammatory microenvironment by the recruitment of lymphocytes. CAFs interacts with epithelial cells to promote the occurrence, growth, invasion and metastasis of gastric cancer cells. In this study, the direct and indirect interaction between CAFs and gastric cancer cells is constructed by three-dimensional cell culture and Transwell. H.pylori is used to infect the both cells to study, in vivo and in vitro, the activation of CAFs, the mechanism of lymphocyte recruitment, the interaction between CAFs and gastric cancer cells and the molecular mechanism of gastric cancer cell growth and metastasis in inflammatory environment. This study will reveal the molecular mechanism of inflammatory microenvironment in gastric cancer induced by H.pylori.

幽门螺杆菌（H.pylori）感染引起的炎性微环境在胃癌的发生发展中有重要作用，而肿瘤相关成纤维细胞(CAFs)构成了肿瘤微环境中主要的细胞群体。前期研究发现H.pylori能主动黏附CAFs，而CAFs能促进胃癌细胞的克隆性生长，用H.pylori诱导的胃癌沙鼠模型伴有进行性大量淋巴细胞的侵润。因此，我们推测H.pylori感染引起的炎性反应，通过直接和间接方式活化成纤维细胞为CAFs， 活化的CAFs分泌各种因子，募集淋巴细胞建立炎性微环境，与上皮细胞相互作用，促进胃癌细胞的发生、生长、侵袭和转移。本项目用三维培养和Transwell小室，构建CAFs与胃癌细胞直接和间接的互作模型，用H.pylori感染，在体内、体外研究炎性环境下CAFs的活化、募集淋巴细胞的机制、CAFs与胃癌细胞的互作、以及促进胃癌细胞生长和转移的分子机制，从炎性微环境的角度揭示H.pylori引起胃癌的分子机制

幽门螺杆菌（H. pylori）感染能破坏胃上皮细胞之间的紧密连接进入细胞，作用于胃上皮细胞、正常成纤维细胞（NFs）及肿瘤相关成纤维细胞（CAFs）等，但是它们之间的互作在胃癌发生发展中的作用尚不清楚。本项目从临床胃癌标本中分离NFs、CAFs及原代胃癌细胞，构建H. pylori、胃癌细胞及NFs/CAFs直接共培养模型及Transwell间接共培养模型，检测细胞因子、NFs活化、胃癌细胞的生长、迁移、侵袭及裸鼠成瘤，研究H. pylori、NFs/CAFs及胃癌细胞的互作促进胃癌的发生发展；建立沉默或过表达Serpin E1的CAFs及胃癌细胞株，研究互作的分子机制并在临床胃癌标本中验证；用H. pylori国际标准株NCTC 11637灌胃感染蒙古沙鼠构建胃癌模型；用H. pylori感染胃癌细胞，研究柠檬酸合酶丙二酰化及线粒体能量代谢。. 取得的主要成果有：（1）NFs与CAFs转录组学鉴定到161个差异表达基因，有利于肿瘤细胞的营养供应和转移；（2）阐明H. pylori、NFs/CAFs及胃癌细胞互作诱导多种细胞因子的释放活化NFs为CAFs，诱导沙鼠胃上皮和人胃癌细胞高表达Serpin E1，促进胃癌细胞的生长、迁移、侵袭及裸鼠成瘤；（3）揭示H. pylori 感染促进CAFs高表达 Serpin E1，诱导人脐静脉内皮细胞VEGFA表达，诱导血管形成促进胃癌的发生发展；（4）证实Serpin E1在人胃癌组织高表达，其高表达与胃癌患者10年总存活率负相关；（5）构建H. pylori GZ7的染色体基因组图；（6）阐明H. pylori抑制去丙二酰化酶SIRT5促进柠檬酸合酶丙二酰化，导致线粒体能量代谢障碍，是H. pylori致病机制。. 总之，研究结果提示靶向Serpin E1通过破坏H. pylori、CAFs及胃癌细胞的互作可能成为治疗胃癌的有效措施。

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