BMP2在慢性传输型便秘肠神经元重塑中作用及其机制的研究

Recently, there has been a significant increase in incidence of slow transit constipation (STC). Previous studies have shown that bone morphogenetic proteins (BMPs) play an important role in the development and differentiation of enteric nervous system(ENS). In our preliminary study, it was found that abnormal expression of BMP2 in colon myenteric ganglia in patients with STC,down-regulation of Ach positive neurons and up-regulation of NOS positive neurons identified a potential role of intestine neuronal plasticity in intestine movement dysfunction in STC. In the present project, intestinal motility and BMP2 changes in myenteric ganglia will be analyzed to determine the relationship between BMP2 and intestine movement dysfunction in STC. A STC animal model in combination with gene knockout mice will be applied to investigate morphological changes and different neurotransmitter secreting cell types to further reveal the correlation between BMPs-Smads pathway and intestine neuronal plasticity. Meanwhile, after myenteric neurons culture or its co-culture with intestinal smooth muscle cells, BMP2, Noggin, iSmad and RNAi will be used to systematically interpret the regulatory network of intestine neuronal plasticity in STC. This study will contribute to the pathogenesis of STC, novel treatment target design and recovery of pathological state in intestinal motility disorders.

近年来慢传输型便秘(STC)发病呈上升趋势。有研究显示，骨形态发生蛋白(BMPs)在肠神经系统(ENS)发生及分化中发挥重要作用，我们前期实验结果发现STC结肠肌间神经节中BMP2存在异常表达，并且Ach阳性神经元表达下调，NOS阳性神经元表达上调，提示肠神经元重塑可能是STC肠道运动功能失调的重要原因。本项目拟检测STC患者肠动力改变、肠肌间神经节BMP2变化，证实BMP2与STC肠道动力异常的相关性；建立STC动物模型，结合基因敲除小鼠，检测肠神经丛形态及不同神经递质分泌细胞类型的改变，揭示BMPs-Smads通路与肠神经元重塑的关系。同时，行体外肌间神经元培养、并与肠平滑肌共培养，应用BMP2、Noggin、抑制性Smad及RNAi进行干预，系统阐明STC肠神经元重塑的网络调控机制。本研究对探求STC的发病机制，寻找治疗的新靶点、逆转肠动力紊乱的病理生理状态，具有重要意义。

近年来慢传输型便秘(STC)发病呈迅速上升趋势。有研究显示，骨形态发生蛋白(BMPs)在肠神经系统(ENS)发生及分化中发挥重要作用。我们前期研究发现肠神经元重塑是STC肠道运动功能失调的重要原因，但其机制未明。本项目通过洛哌丁胺诱导建立了相对稳定的STC小鼠模型，运用改良肠神经铺片技术、免疫荧光等分子生物学方法，发现STC小鼠结肠肌间HUC/D+神经元数目减少，且BMP2、p-Smad1/5/9蛋白表达下调。进行肠道菌群功能富集分析和网络药理学发现薯蓣皂苷（Dioscin）的免疫调节靶点，证实Dioscin通过降低TNFα、IL-6、iNOS炎症因子，调控肠神经元炎症微环境，促进肠肌间巨噬细胞分泌BMP2，激活BMP2/Smads通路，促使HUC/D+神经元数目恢复、神经递质AchE分泌增加，改善肠道动力。同时我们运用16S rRNA和代谢组等多组学研究发现右美沙芬可通过减少肠道菌群丰度，降低肠道菌群多样性，增加短链脂肪酸戊酸、异戊酸的产生，进而调节肠道微生物环境。进一步收集STC结肠组织进行转录组测序，筛选出差异基因NOX1和BMP2，证实右美沙芬不仅可通过抑制NOX1的表达，减少活性氧的产生，抑制氧化应激，导致CD45+白细胞减少，在维持肠道免疫稳态中发挥作用；且右美沙芬可影响小鼠结肠组织BMP2水平，激活BMP2/Smads通路，从而增强肠道动力。本研究深入阐明了BMP2通过Smads通路引起肌间神经元重塑的机制，不仅为STC发病机制研究提供新方法和新思路，而且为肠动力新药创新提供基础和保障。