短链脂肪酸参与肠道菌群与宿主互动致IBS-D内脏高敏感的外周机制及肠安I号方的作用研究

Gut microbiota and their metabolite, short chain fatty acids (SCFAs) play an important role in the pathogenesis of visceral hypersensitivity in diarrhea-predominant irritable bowel syndrome (IBS-D). Our previous work has confirmed that Chang’an I prescription could modulate gut microbiota and improve visceral hypersensitivity in IBS-D patients, however, its mechanism remains largely undefined. In this study, we propose that Chang’an I prescription may regulate gut microbiota, affect SCFAs metabolism in gut and regulate the intestinal permeability through G protein-cAMP-PKA-MAPK signaling pathway mediated by SCFAs and their receptors, GRP41/GPR43. To this end, in the present study, high-throughput sequencing, gas chromatography-mass spectrometry, molecular biology techniques and etc. will be utilized to explore the effect of Chang’an I prescription on regulation of the intestinal permeability through G protein-cAMP-PKA-MAPK signaling pathway via modulation of gut microbiota and SCFAs both in vivo and in vitro. We aim to deeply clarify the peripheral mechanism of the interaction between gut microbiota and host involved in visceral hypersensitivity, as well as the pharmacological action of Chang’an I prescription, which will provide novel theoretical basis and research methods for the clinical therapy of IBS-D.

肠道菌群及其代谢产物短链脂肪酸（SCFAs）在腹泻型肠易激综合征（IBS-D）内脏高敏感发生机制中发挥重要作用。课题组前期研究已证实肠安I号方可以调节IBS-D患者肠道菌群，改善内脏高敏感，但其作用机制不明确。本项目提出肠安I号方可能通过调节肠道菌群，影响肠道SCFAs代谢，在SCFAs及其受体GRP41/GPR43介导下通过G蛋白-cAMP-PKA-MAPK信号通路调节肠黏膜通透性，从而改善内脏高敏感。为此，本项目拟采用高通量测序、气相色谱-质谱联用、分子生物学技术等，从整体、细胞及分子水平研究肠安I号方通过调节肠道菌群及SCFAs，调控G蛋白-cAMP-PKA-MAPK信号通路对肠黏膜通透性的影响，深入阐明肠道菌群与宿主互动参与IBS-D内脏高敏感的外周机制以及肠安I号方的药理作用，为IBS-D的临床治疗提供新的理论依据和研究方法。

肠道菌群及其代谢产物短链脂肪酸（SCFAs）在腹泻型肠易激综合征（IBS-D）内脏高敏感发生机制中发挥重要作用。课题组前期研究已证实肠安I号方可以调节IBS-D患者肠道菌群，改善内脏高敏感，但其作用机制不明确。本项目提出肠安I号方可能通过调节肠道菌群，影响肠道SCFAs代谢，在SCFAs及其受体GRP41/GPR43介导下通过G蛋白-cAMP-PKA-MAPK信号通路调节肠黏膜通透性，从而改善内脏高敏感。为证实以上假说，本研究通过建立IBS-D模型病证结合模型大鼠以及肠安I号方干预，结果显示肠安I号方可以改善模型大鼠内脏高敏感，与影响肠道菌群的结构以及差异菌种的丰度有关，同时肠安I号方影响丙酸、乙酸及丁酸的代谢，同时对己酸、庚酸、辛酸及癸酸的代谢有一定影响。分子机制方面，肠安I号方影响短链脂肪酸相关受体GPR41、GPR43的表达，以及MAPK信号通路中p-JNK的磷酸化水平，从而影响紧密连接蛋白ZO-1、Occludin以及Claudin-1的表达及相关细胞因子IL-6、TNF-α的含量。同时，肠安I号方对TNF-α所致上皮细胞损伤模型具有一定的修复作用，影响细胞TNF-α、IL-6、IL-10的水平，改善黏膜通透性，这与其影响通过影响MAPK信号通路中ERK、JNK及p38蛋白的磷酸化水平，从而进一步调节紧密连接蛋白ZO-1、Occludin及Claudin-1的蛋白水平，影响肠上皮屏障完整性有关。这一研究为进一步深入阐明肠道菌群与宿主互动参与IBS-D内脏高敏感的外周机制以及肠安I号方的药理作用提供了新的思路和依据。