炎症因子 TNF-α促胆囊癌微淋巴管形成的作用及其机制

The occurrence, development and metastasis of gallbladder cancer are closely related to the instigation of chronic inflammation. Lymph node metastasis is the major routes and early events in gallbladder cancer metastases. Lymphangiogenesis is the most important process which influences the metastasis of gallbladder cancer through the lymph node. However, the role of inflammation factors in promoting lymph node metastases of gallbladder cancer has not been clarified. Our previous preliminary research has shown that the inflammatory factor, TNF-alpha, is found in high concentration within the bile of gallbladder cancer. The presence of this cytokine may potentially stimulate VEGF-C secretion by gallbladder cancer cells; which, in conjugation with VEGF-D may promote lymphangiogenesis in gallbladder cancer. The purpose of this study is to explore that the role and mechanism of TNF-alpha in gallbladder cancer lymphangiogenesis and whether this process is mediated by VEGF-C and/or VEGF-D. First, we will aim to address whether TNF-alpha can promote lymphangiogenesis in gallbladder cancer through an in vitro lymphatic tube formation experiment and a transplanted tumor lymphangiogenesis experiment in a murine model. Secondly, we will attempt to confirm whether TNF-alpha can enhance the expression of VEGF-C and/or VEGF-D to promote lymphangiogenesis in gallbladder cancer through mRNA silencing and receptor inhibition of the VEGF-C/VEGF-D experiments. Lastly, we will investigate the related signaling pathways involved in TNF-alpha-mediated upregulation of VEGF-C and/or VEGF-D in gallbladder cancer cells through specific signal transduction blockers acting on tumour cells. These results will help reveal the influencing factors in early metastasis of gallbladder cancer and may lead to new strategies for the prevention and curing of the early metastasis of gallbladder cancer. Furthermore, these results may also shed new light on new drug targets for signaling pathways to further improve the survival rate of gallbladder cancer patients.

胆囊癌的发生和转移与慢性炎症刺激密切相关，淋巴转移是胆囊癌转移的主要途径和早期事件，然而炎症因子在促胆囊癌淋巴转移中发挥的作用尚未阐明。本项目前期研究发现：炎症因子TNF-α在胆囊癌胆汁中高表达；TNF-α可促进体外胆囊癌细胞分泌VEGF-C；VEGF-C和VEGF-D可诱导胆囊癌新生微淋巴管形成。据此，本研究拟探讨TNF-α促胆囊癌微淋巴管形成的作用及其机制：通过人淋巴内皮细胞小管形成实验和裸鼠在体实验，证明TNF-α促进胆囊癌的微淋巴管形成；通过SiRNA沉默VEGF-C/D，抗体阻断VEGFR-3（VEGF-C/D受体）等方式处理细胞，探讨TNF-α是否通过上调VEGF-C和/或VEGF-D促进胆囊癌的微淋巴管形成；通过特异性信号传导阻断剂，揭示TNF-α上调VEGF-C和/或VEGF-D的相关信号通路。研究结果有助于揭示胆囊癌早期转移的影响因素，为胆囊癌早期转移的防治提供新思路。

胆囊癌的发生和转移与慢性炎症刺激密切相关，淋巴转移是胆囊癌转移的主要途径和早期事件，然而炎症因子在促胆囊癌淋巴转移中发挥的作用尚未阐明。本项目前期研究发现炎症因子TNF-α在胆囊癌胆汁中高表达；TNF-α可促进体外胆囊癌细胞分泌VEGF-C；VEGF-C和VEGF-D 可诱导胆囊癌新生微淋巴管形成。据此，本研究探讨了TNF-α促胆囊癌微淋巴管形成的作用及其机制：1）通过人淋巴内皮细胞小管形成实验、人体标本实验和裸鼠在体实验，证明TNF-α促进胆囊癌的微淋巴管形成；2）通过SiRNA沉默VEGF-C/D，抗体阻断VEGFR-3（VEGF-C/D受体）等方式处理细胞，证明TNF-α是通过上调VEGF-C和/或VEGF-D 促进胆囊癌的微淋巴管形成；3）通过特异性信号传导阻断剂，揭示TNF-α上调VEGF-C 和/或VEGF-D 的相关信号通路分别涉及NF-κB和 AP-1。并证明TNF-α通过转录因子NF-κB与VEGF-C启动子相应的位点结合并上调VEGF-C启动子活性，从而上调胆囊癌细胞VEGF-C分泌，并促进胆囊癌微淋巴管生成。TNF-α亦通过“ERK1/2-AP-1”通路增强VEGF-D启动子活性并上调VEGF-D蛋白表达，促进胆囊癌细胞VEGF-D的分泌和胆囊癌微淋巴管生成。该研究结果拓展了胆囊癌的早期影响因素，为胆囊癌早期转移的防治提供新思路。完成研究目标，发表SCI论文5篇，培养研究生5名。