从脂肪细胞命运相关分子网络探讨乌梅丸治疗肥胖症的作用机制

Clinical trial demonstrated that Wu-Mei-Wan　(WMW) could decrease the body weight of obese patients. Our previous studies also showed that WMW can reduce the adipogenesis of white adipose tissue (WAT) and promote the cell proliferation of brown adipose tissue (BAT). It also decreased the expression of Toll-like receptor 3 (TLR3) and can be used for the treatment of obesity, for which the potential mechanism remains unclear. In recent years, it has been found that the regulation of adipocyte directional differentiation play a key role in the treatment of obesity, while the TLR3/IL-6/STAT3 and BMP signaling pathways can mediate the remodeling of adipocyte fate. We put forward the hypothesis that WMW may remodel the adipocyte fate through TLR3/IL-6/STAT3 and BMP signaling pathways　based on what we mentioned above, which indicate WMW may regulate the directional differentiation of adipocyte in the treatment of obesity. In this project, the obese mice induced by high fat diet were studied to explore the effect of WMW on the adipogenesis of WAT and the cell proliferation of BAT. Moreover, cell-based in vitro test will be performed to explore the association between the directional differentiation of adipocyte which is controlled by WMW and the molecular network related to adipocyte fate. It will reveal the scientific connotation by which WMW works on adipose tissue to treat obesity preliminarily and to provide a new insight for WMW as a representative prescription in the treatment of obesity.

临床研究显示乌梅丸能减轻体重，本课题组前期研究提示，乌梅丸减少白色脂肪生成，促进棕色脂肪生成，降低固有免疫模式识别受体TLR3的表达，具有治疗肥胖症的作用，但其机制尚不明确。近年研究发现“调控脂肪细胞定向分化是治疗肥胖症的核心，而TLR3/IL-6/STAT3及BMP信号通路可介导脂肪细胞命运重塑”，本课题以此为切入点，提出“乌梅丸可能通过TLR3/IL-6/STAT3和BMP信号通路重塑脂肪细胞命运，即调控脂肪细胞定向分化治疗肥胖症”的工作假说。本项目拟通过动物实验探讨乌梅丸影响白色脂肪生成，调节棕色脂肪生成及其活性的效应和潜在机制；细胞实验深入研究乌梅丸影响脂肪细胞定向分化与脂肪细胞命运相关分子网络之间的联系。以期初步揭示乌梅丸作用于脂肪组织治疗肥胖症的科学内涵，为以乌梅丸为代表的方剂治疗肥胖症开拓新思路。

本项目运用网络药理学,以高脂饮食诱导的肥胖小鼠、3T3-L1前脂肪细胞及C3H10T1/2多潜能干细胞为研究对象,采用ELISA、RT-qPCR、Western blot、免疫荧光、免疫组化及转录组学测序等实验技术，系统研究了乌梅丸治疗肥胖症的分子机制。网络药理学研究发现,乌梅丸与目前FDA批准治疗肥胖症的药物具有高度相似的作用机制。动物实验发现，乌梅丸通过调控TLR3/IL-6/STAT3信号通路抑制白色脂肪生成；通过调控BMP7/Smad1/5/9信号通路，增强棕色脂肪产热功能，增加机体能量代谢，减轻体重，并改善肥胖相关的代谢紊乱。运用分子对接及细胞实验筛选出乌梅丸中治疗肥胖症的主要活性成分有6-姜酚、山奈酚、熊果酸。在3T3-L1前脂肪细胞成脂分化实验中，6-姜酚通过调节PPARγ、C/EBPα等分子表达，抑制白色脂肪细胞生成。动物实验进一步发现，6-姜酚通过抑制TLR3/IL-6/JAK1/STAT3信号通路，抑制白色脂肪生成，减轻体重，改善胰岛素抵抗及代谢性炎症。在C3H10T1/2细胞实验中，山奈酚与熊果酸均能诱导多潜能干细胞向棕色脂肪细胞定向分化，上调UCP1等蛋白表达，促进线粒体产热。动物实验进一步发现，山奈酚与熊果酸均能有效促进棕色脂肪生成，增加能量代谢，减轻肥胖小鼠体重，其内在作用机制与调控棕色脂肪BMP7、PRDM16信号通路及氧化磷酸化等相关。因此，本研究初步阐明了乌梅丸通过调控脂肪细胞命运相关分子网络治疗肥胖症的机制及其物质基础；为揭示乌梅丸“寒热并用、补泻兼施”治疗肥胖症的现代科学内涵提供了参考。