基于脑-肠-菌群轴建立IBS-D大鼠模型及疏肝健脾法作用机制研究

Diarrhea predominant irritable bowel syndrome(IBS-D) is the most common functional gastrointestinal disorder in our country. The pathophysiology of IBS-D is complex and not completely understood, and there is no ideal animal model. The dysfunction of brain-gut axis and alteration of intestinal flora play an important role in the pathogenesis of IBS-D. The interaction of brain, gut and enteric microbiota form the brain-gut-enteric micobiota axis. Fecal microbiota transplantation was used to treat recurrent Clostridium difficile infection for years. Under this viewpoint, the transplantation of fecal microbiota from patients with IBS-D to animal may lead to diarrhea of animal. This project was designed under the guidance of the concept of brain-gut-enteric micobiota axis and fecal microbiota transplantation. A rat model of IBS-D was established in the method of maternal separation and fecal microbiota transplantation from IBS-D patients. The evaluation of the behavior, character of stools, visceral sensitivity, motility of gastrointestinal tract, the level of brain-gut peptide, action of mast cell and the level of cytokines were did in the rat model. The mechanism of therapy with soothing liver and strengthening spleen was explored in the respect of brain-gut-enteric micobiota axis by metagenomics and so on.

腹泻型肠易激综合征（IBS-D）是我国最常见的功能性胃肠病，其发病机制尚未完全阐明，且缺乏理想的动物模型。脑肠轴功能异常和菌群失调在其发病机制中具有重要的作用，脑、肠和肠道菌群三者之间相互作用，形成了脑-肠-菌群轴。“粪菌移植”可治疗难治性菌群失调相关性疾病，基于“粪菌移植”思路，将IBS-D患者的肠道菌群移植到大鼠体内，有可能复制出IBS-D的腹泻特征。本研究基于脑-肠-菌群轴和“粪菌移植”思路，采用母子分离联合“粪菌移植”法建立IBS-D大鼠模型，并从动物行为学、大便特征、内脏敏感性、胃肠动力、脑肠肽水平、肥大细胞活化和细胞因子水平等方面评价了模型的病理特征。基于该模型，通过分析疏肝健脾法对上述病理特征以及肠道菌群宏基因组学的影响，探讨疏肝健脾法调节IBS-D脑-肠-菌群轴的作用机制。

脑-肠轴功能异常和菌群失调在腹泻型肠易激综合征（IBS-D）发病机制中具有重要的作用，脑、肠和肠道菌群三者之间相互作用，形成了脑-肠-菌群轴。本研究从IBS-D患者粪便中分离培养出优势致病菌，经过探索研究发现将一定浓度的耐药型大肠埃希菌和弗劳地枸橼酸杆菌进行连续灌胃可移植菌群，采用在新生期母子分离应激的基础上联合成年期人源性粪菌悬液灌胃法建立脑-肠-菌群轴异常IBS-D大鼠模型，并对模型进行了全面评价。研究结果显示：（1）与正常大鼠比较，该模型具有内脏敏感性增高、伴腹泻、焦虑样行为特征；（2）血清和远端回肠五羟色胺含量显著增高、远端回肠P物质和血管活性肠肽含量显著降低；（3）近端结肠和远端回肠肿瘤坏死因子α含量显著升高，白介素10含量显著降低；（4）脑皮质和海马辣椒素受体1表达显著升高。（5）与正常大鼠比较，模型组大鼠肠道菌群在门水平脱铁杆菌门消失；在纲水平，模型组丹毒丝菌纲显著降低，脱铁杆菌纲消失；在目水平，模型组支原体目显著升高、丹毒菌丝目显著降低，脱铁杆菌目消失；在科水平，支原体科、韦荣球菌科显著升高，丹毒丝菌科、紫单包菌科、Christensenellaceae科显著降低，优杆菌科、脱铁杆菌科消失；在属水平，疣微菌科-UCG-005及UCG-008属、普氏菌属、Allobaculum属显著降低，瘤胃球菌属2、支原体属、韦荣氏球菌属显著升高。（6）采用疏肝健脾中药复方干预后显示：疏肝健脾中药复方可调整模型大鼠的行为学特征，降低其血清5羟色胺水平及大脑皮质血管活性肠肽含量，可调整模型大鼠肠道菌群的丰度和多样性，从门至属多个水平调整肠道菌群结构，疏肝健脾法具有调节脑-肠-菌群轴的多靶点作用。