基于多组学研究电针对脊髓损伤排便障碍大鼠结肠5-HT系统的微生态调节机制

Intestinal dysbiosis secondary to spinal cord injury affects the functional recovery and the quality of life. Intestinal microflora and metabolites regulate 5-HT system. Combined with previous studies, we hypothesize: acupuncture can regulate intestinal microflora and metabolites, and then regulate the colon 5-HT system by multi-target and multi-channel, and promote intestinal function recovery, which may be an important mechanism to treat defecation disorders in rats with spinal cord injury. This study intends to electroacupuncture (EA) on the defecation disorders rat following spinal cord injury and transplant intestinal microbiota of EA group to model rats. The treatment effect of EA and EA intestinal microbiota was assessed. To measure the influence of EA on intestinal microecology and explore potential therapeutic targets and metabolic pathways of EA, 16SrDNA，Metagenomics high-throughput sequencing，and GC-TOF-MS for metabolomics will be used. Intermediate molecules regulated by intestinal microbiota and various molecules of 5-HT system in colon were determined by ELISA, Real-time PCR, western blot, and immunohistochemistry. The purpose of our project is to reveal the microecological regulation mechanism of 5-HT system controlled by EA, and provide new theoretical and experimental basis for the popularization and application of acupuncture and moxibustion in clinical rehabilitation.

脊髓损伤继发的肠道菌群失调影响患者功能恢复及生活质量；肠道菌群及代谢产物对5-HT系统有调控作用。结合前期研究，我们提出工作假说：针刺通过调节肠道菌群及代谢产物，多靶点、多通路良性调控结肠5-HT系统，促进肠道功能恢复，可能是其治疗脊髓损伤大鼠排便障碍的重要机制之一。拟电针干预脊髓损伤排便障碍大鼠，并以对模型鼠进行电针菌群移植，在对电针和电针菌群的治疗效应进行评价的基础上；采用16SrDNA和宏基因组学高通量测序、气相色谱质谱非靶标代谢组学技术结合生物信息学分析，明确电针对脊髓损伤排便障碍大鼠肠道微生态的调节作用，并探索电针潜在治疗靶点及其代谢通路。采用ELISA、Real time-PCR、Western Blot和免疫组化技术检测肠道菌群调节的中间分子及5-HT系统各分子在结肠中的表达。探讨电针调控5-HT系统的微生态调节机制。从新视角为针灸临床康复的推广应用提供理论和实验依据。

背景：电针能够改善脊髓损伤所致的肠道功能障碍，但是否与其能调控肠道微生态及结肠5-HT 系统有关尚属未知。.研究内容及结果：1.以电针干预脊髓损伤排便障碍大鼠，并对模型鼠进行电针菌群移植，评价电针及菌群移植的治疗效应。结果表明：电针及其菌群干预能缩短首粒黑便排出时间，增加24h排便量，促进SCI大鼠肠道传输功能的恢复，并改善结肠组织形态。2.采用16SrDNA、宏基因组学测序、UHPLC-QTOF-MS非靶标代谢组学检测大鼠粪便并进行生物信息学分析，研究电针对模型大鼠肠道微生态的影响。结果表明：模型大鼠肠道微生物的丰富度、多样性发生了改变，肠道菌群的组成以及变形菌门、梭菌纲、拟杆菌目和多尔氏菌属的数量发生了异常。电针及其菌群移植干预则重塑了肠道菌群。脊髓损伤导致大鼠粪便菌群代谢紊乱，与模型大鼠相比，电针组和菌群移植组儿茶素含量增加。而酚类物质则可能是电针及其菌群移植使SCI大鼠肠道微生物改变与其代谢改变之间有潜在联系的关键物质。此外，还筛选出5个关键差异功能基因及相关代谢通路。3.检测了肠道菌群调节的中间分子及5-HT系统各分子在结肠中的表达。结果表明：模型大鼠结肠5-HT、5HT3AR、5HT4R、SERT和TPH1表达异常，电针及其菌群移植均能良性调节以上分子，并对TLR2和TLR4的表达也发挥良性调节作用。.结论：电针可能通过调节肠道微生物群和代谢产物，调节结肠5-HT系统来改善脊髓损伤大鼠肠道传输功能。.科学意义：本研究明确了电针对脊髓损伤排便障碍大鼠肠道微生态的调节作用，探索了潜在治疗靶点。为针灸治疗本病临床应用的推广提供新的理论和实验依据。同时，电针对肠道菌群及其代谢产物调节作用的研究成果也为未来相关的临床康复应用和基础研究提供了新的方向和思路。