基于“血热理论”探讨清热凉血方调控CD155/TIGIT信号通路抑制T细胞免疫治疗银屑病的分子机制

It is generally accepted that syndrome of heat toxicity due to heat stagnating in blood is the core pathogenesis of psoriasis. Heat-clearing and blood-cooling formula (HCBCF), which is on the principle of clearing heat and removing toxicity, and cooling blood and invigorating circulation, has a good effect on psoriasis with blood heat syndrome. However, the mechanisms and effect targets of HCBCF remain unclear. T-cell immunoglobulin and immunoreceptor tyrosine-based inhibition motif domain (TIGIT) can inhibit the differentiation of CD4+T cells into Th1/Th17 cells, thus inhibiting the activation of T cells and improving the excessive proliferation of psoriatic epidermis. Our previous research has found that HCBCF could regulate CD4+T lymphocyte proliferation through affecting Notch signaling pathway. At present, there is no study on molecular mechanism of HCBCF or the other Chinese herbs for psoriasis via CD155/TIGIT signal pathway. We hypothesized that HCBCF could inhibit the differentiation of CD4+T cells into Th1/Th17 via activating the CD155/TIGIT signaling pathway, thus inhibiting the proliferation and activation of T cells. In this project, both animal studies and cell culture experiments will be performed by using morphology techniques, HE staining, immunohistochemistry, RT-PCR and so on. The purpose of the project is to explore the molecular mechanism of HCBCF to improve psoriasis by regulating the CD155/TIGIT signaling pathway to inhibit the proliferation and activation of T cells, and to provide new ideas for the research and application of traditional Chinese medicine in the treatment of psoriasis.

血热内蕴、郁久化毒被广泛认可是银屑病的核心病机，以清热解毒、凉血活血为治法的“清热凉血方”治疗银屑病血热证疗效确切，其作用靶点和具体机制尚不明确。TIGIT能抑制CD4+T细胞向Th1/Th17细胞分化而抑制T细胞活化，改善银屑病表皮过度增殖。我们前期研究发现：“清热凉血方”可通过影响Notch信号通路调节CD4+T淋巴细胞增殖。目前，尚无“清热凉血方”或其他中药通过CD155/TIGIT信号通路调控银屑病的分子机制研究。我们推测：清热凉血方可通过激活CD155/TIGIT信号通路，抑制CD4+T细胞向Th1/Th17分化，进而抑制T细胞增殖活化。本项目拟从动物研究和细胞培养两层次，采用形态学、HE染色、免疫组化、RT-PCR等技术，探讨清热凉血方通过调控CD155/TIGIT信号通路抑制T细胞增殖活化而改善银屑病的分子机制，为中医药治疗银屑病的研究和应用提供新思路。

银屑病发生发展与T细胞过度活化密切相关，T细胞表面共抑制分子在T细胞活化过程中起关键作用。TIGIT是一种免疫毒性低于CTLA-4、PD-1等的共抑制分子，可以通过免疫应答抑制T细胞的活化增殖。本项目通过建立咪喹莫特银屑病小鼠模型，提取原代银屑病样小鼠的脾脏T淋巴细胞，验证TIGIT在银屑病发病中的作用及清热凉血方是否可以激活CD155/TIGIT信号通路抑制T细胞向Th1/Th17的分化治疗银屑病。研究结果显示：（1）TIGIT在银屑病小鼠外周血及脾脏T细胞中呈低表达状态，说明TIGIT信号通路与负性调节银屑病T细胞活化增殖密切相关。（2）清热凉血方对TIGIT的促进效果与激活剂CD155类似，可促进CD155/TIGIT信号通路的激活，进而抑制CD4+T细胞的增殖，降低细胞因子IFN-γ、IL-17A的产生，从而发挥治疗银屑病的作用。因此，CD155/TIGIT信号通路可能参与银屑病的发病过程，其中TIGIT可能成为银屑病治疗的有效靶点。清热凉血方是治疗血热型银屑病的有效方剂，针对血热型病因病机，治以清热解毒，凉血活血，并通过激活CD155/TIGIT信号通路抑制T细胞活化，继而抑制炎症因子的表达发挥治疗银屑病的作用。