ROS-HIF-1α-AQP3-ROS环路介导胃癌细胞分泌TGF-β1上调CAFs表达HAPLN1促进胃癌侵袭转移的机制研究

Tumor invasion and metastasis have a major effect on the prognosis of gastric carcinoma. We have demonstrated that aquaporin 3 (AQP3) promotes gastric cancer invasion and metastasis via inducing epithelial-to-mesenchymal transition and the stem-like properties previously. More recently, our studies indicate that H. pylori infection can promote AQP3 expression regulated via the ROS-hypoxia-inducible factor 1α (HIF-1α) axis, and that the ROS-HIF-1α-AQP3-ROS loop results in exacerbating this molecular process. Importantly, the loop promotes the secretion of TGF-β1 in gastric cancer cells. The expression of HAPLN1 in gastric cancer associated fibroblasts (CAFs) increased significantly with regarding to that in non-cancer fibroblasts, and TGF-β1 treatment augmented HAPLN1 production in CAFs. Therefore, we put forward the hypothesis: TGF-β1 secreted by tumor cells via ROS-HIF-1α-AQP3-ROS loop up-regulates HAPLN1 expression of CAFs to remodel the extracellular matrix (ECM), and further to promote the invasion and metastasis of gastric carcinoma under the condition of H. pylori infection. To verify this hypothesis, a series of studies, including clinicopathological investigation, in vitro and in vivo experiments (models of mouse and zebra fish), are arranged to be conducted to probe the mechanisms underlying cell interaction between tumor cells and CAFs via TGF-β1/Smad/HAPLN1 to promote tumor invasion and metastasis. This study will help to further elucidate the molecular pathomechanism of cell interactions inside tumor microenvironment in facilitating gastric cancer progression, and provides novel strategies for the prevention and treatment of gastric cancinoma.

侵袭转移严重影响胃癌预后。我们研究表明水通道蛋白3（AQP3）通过调节胃癌细胞EMT及干细胞干性获得促进胃癌侵袭转移。前期研究提示，H. pylori感染通过ROS-HIF-1α-AQP3-ROS环路增加胃癌细胞AQP3表达，并促进TGF-β1分泌；胃癌CAFs的HAPLN1表达显著高于NFs，而TGF-β1上调CAFs的HAPLN1表达。据此我们提出假说：H. pylori感染通过上述环路介导胃癌细胞分泌TGF-β1，进而经TGF-β1/Smad途径使CAFs分泌HAPLN1增加，重构细胞外基质，促进胃癌侵袭转移。本课题拟从细胞分子、临床标本组织及动物模型等多层次探讨H. pylori感染促进胃癌细胞分泌TGF-β1的机制；确定TGF-β1调控CAFs分泌HAPLN1；揭示HAPLN1促进胃癌侵袭转移机制。本研究将从胃癌细胞与其CAFs相互作用探讨胃癌侵袭转移机制，为胃癌防治提供新思路。

幽门螺杆菌（H. pylori）感染可诱导缺氧微环境形成，而缺氧微环境影响胃癌细胞与肿瘤相关成纤维细胞（CAFs）相互作用促进肿瘤侵袭转移。本项目旨在研究H. pylori感染诱导缺氧微环境下胃癌细胞与CAFs的相互作用促进胃癌侵袭转移的机制。.本项目从细胞分子、临床标本检测及动物模型等多层次探讨（1）H. pylori感染促进胃癌细胞分泌TGF-β1的机制、（2）TGF-β1调控CAFs分泌HAPLN1的途径及（3）HAPLN1重构细胞外基质促进胃癌侵袭转移的机制。.主要结果：.（1）H. pylori感染可激活胃癌细胞ROS-HIF-1α-AQP3-ROS环路并通过HIF-1α促进胃癌细胞TGF-β1的转录与表达。.（2）临床标本研究证实H. pylori感染与胃癌细胞中ROS-HIF-1α-AQP3-ROS环路激活及TGF-β1的分泌显著正相关，并与患者预后呈负相关。.（3）小鼠H. pylori感染模型研究证实H. pylori感染后胃黏膜细胞ROS-HIF-1α-AQP3-ROS环路被激活及TGF-β1分泌显著增加。.（4）体外研究证实胃癌细胞分泌的TGF-β1作用于CAFs后，经TGF-β1/Smad途径促进CAFs进一步活化，并促进HAPLN1分泌增加；3D侵袭实验证实HAPLN1表达能够重塑细胞外基质，促进胃癌侵袭和转移。.（5）临床标本检测证实HAPLN1在胃癌组织中高表达，且主要位于α-SMA阳性的间质细胞中；胃癌组织中TGF-β1表达与HAPLN1表达显著正相关；双光子实验证实HAPLN1表达与细胞外基质重构显著正相关，临床数据表明与患者预后呈负相关。.（6）裸鼠移植瘤模型，证实胃癌细胞分泌TGF-β1，经TGF-β1/Smad途径作用于CAFs，促进HAPLN1分泌；双光子实验证实细胞外基质发生重构，促进了胃癌侵袭和转移。.结论和科学意义：.H. pylori感染通过ROS-HIF-1α-AQP3-ROS环路介导胃癌细胞分泌TGF-β1，经TGF-β1/Smad途径促进CAFs分泌HAPLN1，重构细胞外基质，促进胃癌侵袭转移。通过本研究有助于阐明胃癌微环境中胃癌细胞与其CAFs相互作用促进胃癌侵袭转移的机制，为胃癌防治提供新策略。