西红花苷和绿原酸组合通过TGR5/FXR信号系统调控肥胖症肝、肠细胞胆汁酸代谢

Obesity has increasingly become a major global health issue, and there is a lack of effective and safe drug for the treatment of this disease nowadays. Qsymia, which was approved by FDA in 2012, describes the trend of future development of the anti-obesity combination agents. The abnormalities of bile acid metabolism plays a critical role in the etiology of obesity, and G-protein-coupled bile acid receptor (TGR5) and Farnesoid X receptor (FXR) signaling have a key role in regulating those processes. Our previous research found that the combination of the extracts of Gardenia jasminoides and the leaf of Eucommia ulmoides Oliver had anti-obesity effects, which were closely associated with bile acid metabolism. Crocin and Chlorogenic acid are the principal active constituents in those extracts, which have synergistic effect. We hypothesized that the synergistic anti-obesity effect of crocin and chlorogenic acid would be realized by TGR5/FXR signal pathway to modulate bile acid metabolism in hepatocytes and enterocytes and anti-inflammatory. In this study: 1) The anti-obesity effects of the combination of Crocin and Chlorogenic acid will be examined in vivo using high-fat diet (HFD)-induced alimentary obesity of SD rats and monosodium glutamate (MSG)-treated metabolism dysfunctional obesity of C57BL/6 mice; the possible influences of the drug combinations for bile acid metabolism will be studied based on metabonomics as well as inflammatory markers; 2) To investigate the regulating effects on bile acid transporters and inflammatory markers and chemokines of these drugs by TGR5 and FXR, some tool agents and siRNA gene knockdown and plasmid infection technique will be used in two cell models stimulated by bile salts: the HepG2 human hepatoma cells and the murine intestinal STC-1 cells. This study is supposed to provide theoretical and experimental bases for the development of anti-obesity drugs.

肥胖是困扰人类的全球性疾病，尚缺乏安全有效的治疗药物，2012年FDA批准的Qsymia为协同减肥药物的研究指明了方向。胆汁酸激活G蛋白偶联膜受体(TGR5)和法尼酯X受体(FXR)拮抗肝脏、肠道炎症，调节能量平衡。我们前期研究发现栀子和杜仲叶提取物影响胆汁酸代谢协同减肥，而西红花苷和绿原酸作为二者主要有效成分具有协同效应。我们提出假说：西红花苷和绿原酸协同预防肥胖，可能通过FXR和TGR5信号调控肝肠胆汁酸代谢拮抗炎症而实现。本课题拟以高脂饮食诱导肥胖SD大鼠和谷氨酸钠诱导C57BL/6J肥胖小鼠，代谢组学方法观察西红花苷和绿原酸组合预防肥胖症干预胆汁酸代谢及炎症因子表达；以胆盐刺激人肝癌HepG2和小鼠小肠内分泌STC-1细胞，利用工具药、siRNA干扰及过表达质粒转染等方法，探讨药物通过TGR5和FXR信号系统对胆汁酸转运体、炎症及趋化因子的表达调控。本研究可为减肥药物开发提供依据。

肥胖是困扰人类的全球性疾病，尚缺乏安全有效治疗药物。胆汁酸激活G蛋白偶联膜受体(TGR5)和法尼酯X受体(FXR)拮抗肝脏、肠道炎症，调节能量平衡。前期发现栀子和杜仲叶提取物影响胆汁酸代谢协同减肥，西红花苷(CRO)和绿原酸(CGA)是二者有效成分。据此以谷氨酸钠诱导C57BL/6J肥胖小鼠、高脂饮食诱导SD肥胖大鼠，观察CRO和CGA预防肥胖的药效，抗炎及对糖脂、胆汁酸代谢的影响；并以CDCA刺激人肝癌HepG2和小鼠肠内分泌STC-1细胞，采用激动剂和阻断剂以及过表达质粒和shRNA转染等方法，探讨药物通过FXR、TGR5对胆汁酸转运体、炎症及趋化因子表达调控的机制。结果表明CRO和CGA单剂量和联合给药，均显著改善肥胖动物体重体型和血脂血糖，增加胆汁流量，促进脂质和胆汁酸排出，抑制内脏脂质沉积，抗炎维护肝脏和肠道的正常形态和功能，联合用药效果优于单剂量。CRO和CGA预防动物肥胖与改善胰岛素抵抗、抑制慢性炎症状态、改善肝脏和回肠FXR、TGR5和胆汁酸转运体的基因和蛋白表达异常，调控胆汁酸亚组分含量，影响胆汁酸肝肠代谢有关。CRO和CGA协同促进HepG2细胞自噬，降低胆盐毒性，显著抑制肝、肠细胞内脂质和胆汁酸的异常积聚及炎症和趋化因子的分泌。CRO和CGA协同激动胆汁酸受体FXR、TGR5调控HepG2细胞SHP-2/CYP7A1和STC-1细胞SHP-2/FGF15/FGFR4信号通路，促进胆汁酸排出并抑制胆汁酸吸收转运体的基因和/或蛋白表达，调节肝肠循环维持胆汁酸稳态，同时证明CRO和CGA激动作用强于FXR、TGR5的经典激动剂GW4064、INT777。综上本项目获得CRO+CGR通过FXR和TGR5信号调控肝肠胆汁酸代谢，抗炎、改善糖脂代谢而协同预防肥胖的直接证据，为阐明减肥机制提供新思路，推进其作为减肥药物的研究。