基于代谢组学的小儿泄泻脾虚证证候及效应机制研究

Diarrhea is one of the common digestive diseases in children. Persistent and chronic diarrhea caused by a variety of factors in children may have the characteristics of spleen deficiency syndrome. In our previous studies, it has been showed that Wen Yun particle is clinically effective for infantile diarrhea with spleen deficiency syndrome. Regulation of intestinal flora is one of the possible mechanisms. However, the biological basis of the syndrome and the mechanisms of Wen Yun particle remain unclear.In this study, Liquid Chromatography-Mass Spectrometry (LC-MS) and Gas Chromatography - Mass Spectrometry (GC-MS) detection techniques, integrating with chemometric analysis techniques are applied to explore the metabolites of diarrhea with spleen deficiency syndrome and mechanisms of Wen Yun particle. Firstly, urine and faeces samples of clinicians are analyzed by GC-MS to describe the distinctive metabolic profiles of spleen deficiency syndrome of diarrhea before and after treatment of Wen Yun particle. The biomarker extraction are analyzed based on the multivariant analysis. Then by metabolic network construction, the scientific essence of syndrome is demonstrated. Finally, the microscopic mechanism of Wen Yun particle is explained. The findings are expected to give insights into scientific essence of diarrhea with spleen deficiency syndrome and the mechanisms of its effective prescription.

腹泻是小儿时期常见的消化系统疾病之一。各种因素导致的小儿迁延性、慢性腹泻往往具有脾虚泻的证候特点。前期研究基础表明：温运颗粒是治疗小儿泄泻脾虚证的临床有效方药；调节肠道菌群是其可能的作用机制之一。为实现泄泻脾虚证证候客观量化表征，探讨其临床有效方药的可能机制，本项目采用液相色谱-质谱联用检测技术、气相色谱-质谱联用检测技术和化学计量学分析研究技术，研究小儿泄泻脾虚证的代谢组学特征及有效方药温运颗粒对其干预后产生的标志性代谢产物。通过研究泄泻脾虚证患儿体液（尿液和粪便）中代谢网络的变化特征，建立基于代谢成分表达差异的小儿泄泻脾虚证的特征谱库，探讨脾虚泻的发病机制。通过研究温运颗粒对泄泻脾虚证干预后所产生的标志性代谢产物，推断可能的代谢途径，为泄泻脾虚证有效方药的研制提供依据。

研究利用液相色谱-质谱联用技术（LC-MS）、气相色谱-质谱联用技术（GC-MS）分析已采集的尿样、粪样，运用化学计量学方法如主成分分析、偏最小二乘法及非线性数学模型处理数据，筛选证候归属的差异性代谢物，分析相关代谢通路，探讨泄泻脾虚证的发病机制及疗效机制。研究结果表明：脾虚泻和正常患儿的尿液和粪便差异性代谢物共同参与的代谢通路：精氨酸和脯氨酸代谢、β-丙氨酸代谢、甘氨酸，丝氨酸和苏氨酸代谢、氮代谢、苯丙氨酸代谢。同时提出全国名老中医汪受传临床经验方温运颗粒治疗脾虚泻的可能机制为：通过甘氨酸，丝氨酸和苏氨酸代谢、精氨酸和脯氨酸代谢通路改善肠粘膜代谢能量供给、促进粘膜修复；通过调节短链脂肪酸和色氨酸代谢通路进而调节肠道菌群平衡、改善肠道功能，发挥运脾止泻功效。本研究结果为临床验方温运颗粒治疗脾虚泻的机制阐释提供了一定的思路，为今后进一步开展分子机制探讨提供方向。