Jiang Hailun, Zeng Li, Dong Xueqi, Guo Shuilong, Xing Jianguo, Li Zhuorong, Liu Rui
Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China.
Department of Cardiology, Fuwai Hospital, National Center for Cardiovascular Disease, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China.
Front Pharmacol. 2020 Mar 4;11:205. doi: 10.3389/fphar.2020.00205. eCollection 2020.
Human pharyngeal squamous cell carcinoma is highly invasive and proliferative, and exhibits an extremely low 5-year survival rate due to poor understanding of the underlying pathogenic mechanisms, and lack of efficient treatment. It has been shown that the immunosuppressive microenvironment created by tumor cells impairs the immune response against tumor progression, thereby affecting the prognosis for tumor patients. Thus, to improve therapeutic efficacy, it is critical to identify novel drugs with immunoinflammatory modulatory properties to treat tumor immune evasion. Tilianin, the main ingredient of total flavonoids extracted from L., has multiple biological functions, including cardiovascular protective effects, anti-tumor effects, and anti-inflammatory effects. In the present study, the suppressive effects of tilianin on human pharyngeal squamous cell carcinoma were investigated and the underlying mechanisms in regulating the tumor immunosuppressive microenvironment were explored. The cytotoxicity of tilianin on FaDu cells was determined by CCK-8 and clone formation assays. Moreover, the levels of toll-like receptor 4 (TLR4) signaling transduction and apoptotic pathways were determined by immunocytochemical, biochemical, and molecular biological technologies. In addition, the maturation of dendritic cells (DCs) that were co-cultured in supernatant of FaDu cells was evaluated by flow cytometry to investigate alterations in immune system function. For mechanistic exploration, TLR4 siRNA, p38 siRNA, c-Jun N-terminal kinase (JNK) siRNA, and p65 siRNA were used as loss-of-function target evaluation of tilianin therapy. Combined, these results showed that tilianin treatment increased cytotoxicity as well as the apoptotic population of FaDu cells in a dose-dependent manner. Furthermore, tilianin treatment decreased the level of anti-apoptotic markers Bcl-2 and Bcl-xL, increased the level of apoptotic factors Bad and Bax, and stimulated cytochrome release, caspase-3 and poly ADP ribose polymerase (PARP) activation in FaDu cells. Furthermore, our findings indicated that tilianin treatment activated TLR4/p38/JNK/NF-κB signaling pathways and increased the release of inflammatory cytokines. This promoted the maturation of DCs to enhance immune system function in the tumor microenvironment. Moreover, the effects of tilianin on immune system function were abolished by TLR4 siRNA and p65 siRNA. In conclusion, these findings suggested that tilianin may be of immunotherapeutic value for inhibiting human pharyngeal squamous cell carcinoma.
人咽鳞状细胞癌具有高度侵袭性和增殖性，由于对其潜在致病机制了解不足且缺乏有效治疗方法，其5年生存率极低。研究表明，肿瘤细胞产生的免疫抑制微环境会损害针对肿瘤进展的免疫反应，从而影响肿瘤患者的预后。因此，为提高治疗效果，识别具有免疫炎症调节特性的新型药物以治疗肿瘤免疫逃逸至关重要。田基黄苷是从地锦草中提取的总黄酮的主要成分，具有多种生物学功能，包括心血管保护作用、抗肿瘤作用和抗炎作用。在本研究中，研究了田基黄苷对人咽鳞状细胞癌的抑制作用，并探讨了其调节肿瘤免疫抑制微环境的潜在机制。通过CCK-8和克隆形成试验测定田基黄苷对FaDu细胞的细胞毒性。此外，采用免疫细胞化学、生物化学和分子生物学技术测定Toll样受体4（TLR4）信号转导和凋亡途径的水平。此外，通过流式细胞术评估在FaDu细胞上清液中共培养的树突状细胞（DC）的成熟情况，以研究免疫系统功能的变化。为进行机制探索，使用TLR4 siRNA、p38 siRNA、c-Jun氨基末端激酶（JNK）siRNA和p65 siRNA作为田基黄苷治疗的功能丧失靶点评估。综合来看，这些结果表明田基黄苷治疗以剂量依赖方式增加了FaDu细胞的细胞毒性以及凋亡细胞数量。此外，田基黄苷治疗降低了抗凋亡标志物Bcl-2和Bcl-xL的水平，增加了凋亡因子Bad和Bax的水平，并刺激了FaDu细胞中细胞色素c的释放、caspase-3和聚ADP核糖聚合酶（PARP）的激活。此外，我们的研究结果表明，田基黄苷治疗激活了TLR4/p38/JNK/NF-κB信号通路，并增加了炎性细胞因子的释放。这促进了DC的成熟，以增强肿瘤微环境中的免疫系统功能。此外，TLR4 siRNA和p65 siRNA消除了田基黄苷对免疫系统功能的影响。总之，这些发现表明田基黄苷在抑制人咽鳞状细胞癌方面可能具有免疫治疗价值。
