基于Th17/Treg平衡探索木香烃内酯抗溃疡性结肠炎的作用机制

Ulcerative colitis (UC) is a chronic and non-specific intestinal inflammatory disease that lacks effective therapeutic drugs in clinic. The imbalance of Th17/Treg plays a key role in the pathogenesis of UC, and restoring its balance through reducing the Th17 cell or elevating the Treg cell percentages is an important strategy for the treatment of UC. Therefore, looking for anti-UC drug and clarifying its mechanism on the basis of Th17/Treg balance will have more important scientific research value and clinical significance. Our previous studies demonstrated that costunolide possessed a good anti-UC effect and were able to reverse the Th17/Treg imbalance in UC mice. Exploratory studies indicated that costunolide significantly decreased the H3K4me3 and increased the H3K27me3 protein expressions in colons of UC mice. In addition, costunolide can reduce the lactate levels in colons of UC mice. Based on these results, we propose a scientific hypothesis that “costunolide inhibits the process of glycolysis and then influences the histone methylation modification to restore the Th17/Treg balance, which will eventually restrain the development of UC”. In this project, we intend to use RNA interference and other techniques in combination with in vitro experiments to reveal the molecular mechanism of the anti-UC effect exerted by costunolide. The successful implementation of this study will provide a scientific theoretical basis for the development of anti-UC drugs from radices saussureae.

溃疡性结肠炎（UC）是一种慢性非特异性肠道炎症性疾病，临床缺乏有效的治疗药物。Th17/Treg免疫失衡在UC发病中占据关键地位，通过下调Th17或上调Treg细胞比例来恢复Th17/Treg平衡是治疗UC的重要策略。因此，针对Th17/Treg平衡寻找有效的抗UC药物并阐明其作用机制，将具有更重要的科研价值和临床意义。我们前期研究发现木香烃内酯具有良好的抗UC作用，能够扭转UC小鼠的Th17/Treg失衡。探索性研究发现木香烃内酯显著下调UC小鼠结肠组织H3K4me3的表达，上调H3K27me3的表达，降低乳酸水平。基于此，我们提出研究假说：木香烃内酯可能通过抑制糖酵解途径，调节组蛋白甲基化修饰，恢复Th17/Treg平衡，最终达到抗UC的作用。本项目拟采用RNA干扰等技术，结合体外细胞实验，重点开展木香烃内酯抗UC作用的分子机制研究，为从木香药材中开发抗UC药物提供理论依据。

溃疡性结肠炎（ulcerative colitis, UC）是一种慢性特发性病变部位主要在结肠的炎症性疾病。木香的主要活性成分木香烃内酯（Costunolide）被证实具有抗炎和免疫调节活性。本项目首先考察了costunolide对葡聚糖硫酸钠和2, 4, 6-三硝基苯磺酸诱导小鼠UC的影响。结果显示，costunolide（25, 50 mg/kg）灌胃给药可显著降低结肠炎小鼠的疾病活动指数评分，提高结肠炎小鼠生存率，抑制结肠长度缩短，下调结肠组织髓过氧化物酶活力，改善结肠组织病理损伤并降低结肠组织前炎因子TNF-α、IL-1β和IL-6的水平。此外，costunolide（12.5, 25, 50 mg/kg）对健康小鼠体重、外周免疫细胞分型、各组织器官重量及病理损伤均无明显影响。提示，costunolide是一个有效且安全的抗UC药物。Th17/Treg平衡被证实参与UC的发生发展过程。Costunolide（25, 50 mg/kg）能够恢复脾脏、肠系膜淋巴结和结肠组织中Th17/Treg细胞平衡，表现为Th17细胞比例减少、Rorc, Il17a mRNA表达降低和Treg细胞比例微弱增加、Foxp3, Il10 mRNA表达微弱升高。体外研究发现，costunolide（1, 2 μM）直接抑制Th17细胞分化而不影响树突状细胞成熟。深入机制探索发现，costunolide促进脯氨酰羟化酶-2介导的HIF-1α泛素化-蛋白酶体降解，抑制糖酵解途径，从而减少α-酮戊二酸生成，抑制组蛋白去甲基化酶JMJD3活性，增加Rorc启动子区域H3K27me3富集。上述过程最终阻断Rorc转录过程，减少Th17细胞分化。本研究揭示了costunolide恢复Th17/Treg平衡从而发挥抗UC作用的靶点及分子机制，裨益costunolide和其它中药单体对UC的临床治疗。