风湿宁粪菌移植物干预类风湿关节炎的多组学比较研究  

A multiomics comparative study on the intervention of fecal microbiota transplants of Fengshining on rheumatoid arthritis

作　　者：马艳苗 郝佳瑨 刘明燃 宋博 魏福霞 彭涛 MA Yan-miao;HAO Jia-jin;LIU Ming-ran;SONG Bo;WEI Fu-xia;PENG Tao(Shanxi University of Chinese Medicine,Taiyuan 030619,China;Shenyang Pharmaceutical University,Shenyang 110016,China;Shanxi Hospital of Integrated Traditional and Western Medicine,Taiyuan 030013,China)

机构地区：[1]山西中医药大学,山西太原030619 [2]沈阳药科大学,辽宁沈阳110016 [3]山西省中西医结合医院,山西太原030013

出　　处：《药学学报》2023年第7期1931-1939,共9页Acta Pharmaceutica Sinica

基　　金：国家自然科学基金资助项目(82104731);山西中医药大学2022年度科技创新团队(2022TD2005);山西中医药大学2021年研究生创新创业项目(2021CX012);2021年度山西省研究生教育创新计划项目(2021Y722).

摘　　要：运用超高效液相色谱-四极杆-静电场轨道阱高分辨质谱法(ultra-performance liquid chromatographyquadrupole/electrostatic field obitrap high-resolution mass spectrometry,UHPLC-Q-Exactive Orbitrap-MS)及宏基因组学技术,探讨风湿宁粪菌移植干预类风湿关节炎的作用机制。所有动物福利和实验过程均遵循山西中医药大学医学伦理委员会的规定。将大鼠随机分为空白组、模型组、粪菌移植组、雷公藤多苷组,建立大鼠胶原诱导关节炎(collagen induced arthritis,CIA)模型。评估大鼠体重变化、足跖病变情况,采用液质联用、宏基因组技术对各组大鼠血清进行差异代谢物、菌群分析,Western blot检测Toll样受体4(Toll-like receptors,TLR4)、TLR4启动髓系分化初级反应蛋白88(myeloid differentiation factor88,MyD88)和核因子-κB p65(nuclear factor of kappa B,NF-κB p65)的蛋白表达量。从大鼠血清中共筛选出13种差异代谢物:花生四烯酸、二十二碳六烯酸、13S-羟基十八碳二烯酸和L-苯丙氨酸等,经通路富集筛选出3条代谢通路:苯丙氨酸、酪氨酸和色氨酸生物合成、苯丙氨酸代谢,花生四烯酸代谢。宏基因组分析发现CIA组g_Bacteroides、g_Prevotella、p_Actinobacteria丰度较高;FMT-F组c_Clostridia、g_Akkermansia、s_Akkermansia_muciniphila丰度较高。层次聚类热图显示,阿克曼菌(Akkermansia)与L-苯丙氨酸呈负相关,与二十二碳六烯酸呈正相关;普雷沃菌(Prevotella)与L-苯丙氨酸呈正相关。FMT-F能够明显抑制大鼠滑膜TLR4、MyD88和p65蛋白的表达(P<0.01)。风湿宁粪菌移植治疗类风湿关节炎可能与其通过Akkermansia、Prevotella等微生物干预苯丙氨酸、花生四烯酸代谢,抑制TLR4/MyD88/NF-κB通路,发挥抗炎效应有关。The study aims to investigate the mechanism of Fengshining fecal microbiota transplants in the intervention of rheumatoid arthritis by ultra-performance liquid chromatography-quadrupole/electrostatic field obitrap high-resolution mass spectrometry(UHPLC-Q-Exactive Orbitrap-MS).All animal welfare and experimental procedures followed the regulations of the Medical Ethics Committee of Shanxi University of Chinese medicine.The rats were randomly divided into normal group,model group,fecal microbiota transplantation group and Tripterygium wilfordii polyglycoside group,and the collagen induced arthritis(CIA)was established.The changes of body weight and metatarpodal lesions of rats were evaluated.The serum of rats in each group was analyzed by liquid chromatography-mass spectrometry and metagenomic technology for differential metabolites and microflora.The protein expression levels of Toll-like receptors(TLR4),myeloid differentiation factor 88(MyD88)and nuclear factor of kappa B(NF-κB p65)were detected by Western blot.A total of 13 different metabolites,including arachidonic acid,docosahexaenoic acid,13S-hydroxyoctadecanodienoic acid and L-phenylalanine were screened from serum.Three metabolic pathways,including phenylalanine,tyrosine and tryptophan biosynthesis,phenylalanine metabolism and arachidonic acid metabolism were identified through pathway enrichment.Metagenomic analysis showed that the abundance of g_Bacteroides,g_Prevotella and p_Actinobacteria in CIA group was higher.The abundance of c_Clostridia,g_Akkermansia and s_Akkermansia_muciniphila in fecal microbiota transplantation group is higher.The hierarchical cluster heat map showed that Akkermansia was negatively correlated with L-phenylalanine;while positively correlated with docosahexaenoic acid.Prevotella was positively correlated with L-phenylalanine.Fecal microbiota transplantation group could significantly inhibit the expression of TLR4,MyD88 and p65 proteins in the synovium of rats(P<0.01).The anti-rheumatoid arthritis effects of fecal microbiota tr

关 键 词：风湿宁 粪菌移植 肠道菌群 类风湿关节炎 代谢组学 

分 类 号：R917[医药卫生—药物分析学]