背景:绞股蓝(Thunb。)牧野(G.绞股蓝)是一种东方草药,有文献记载可以治疗许多疾病,包括肥胖,高脂血症,代谢综合征和衰老。然而,绞股蓝的抗肥胖机制仍然知之甚少。
目的:通过非靶向代谢组学研究揭示绞股蓝提取物(GPE)在高脂饮食(HFD)诱导的肥胖小鼠中的抗肥胖机制,实时定量PCR(RT-qPCR),和免疫组织化学实验。此外,通过LC-MS/MS和分子对接方法初步鉴定活性成分。
方法:采用乙醇回流法制备GPE,用HP-20大孔树脂纯化。通过液相色谱-质谱(LC-MS)系统鉴定GPE的组分。将42只C57BL/6J小鼠随机均分为6组,每组7只小鼠:对照组,肥胖模型组,Beinaglutide组(阳性对照),GPE低,中等,和高剂量组(50mg/kg,100mg/kg,和200mg/kg的80%乙醇提取物)。体重,肝脏重量,血糖,血脂,并评估肝脏组织病理学变化。非靶向代谢组学用于表征GPE治疗后肥胖小鼠的代谢变化。使用实时定量PCR(RT-qPCR)和免疫组织化学实验验证与差异代谢物相关的基因的表达。通过分子对接方法初步鉴定了GPE具有抗肥胖作用的成分。
结果:在GPE中鉴定出总共17种化合物。GPE显著降低了体重,总胆固醇(TC)和低密度脂蛋白胆固醇(LDL-C)在肥胖小鼠和减少肝脏重量和肝脏脂肪变性。血清代谢组学鉴定出33种与肥胖小鼠GPE治疗相关的潜在生物标志物,主要与色氨酸代谢有关。GPE处理下调Slc6a19和Tph1的表达并上调Ucp1的表达。分子对接说明了20(R)-人参皂苷Rg3、五倍子皂苷I、DamulinB,绞股蓝苷L,油菜花苷B,在GPE中鉴定出的Tricin7-新橙皮苷与Tph1表现出良好的相互作用。
结论:绞股蓝提取物可以通过Slc6a19/Tph1途径抑制色氨酸的吸收及其向5-HT的转化,上调Ucp1的表达,从而促进棕色脂肪组织的产热,促进减肥,减轻脂肪肝的症状。三萜类化合物如AraliasaponinI,在GPE中确定,可能是Tph1的潜在抑制剂,并负责抗肥胖活性。
BACKGROUND: Gynostemma pentaphyllum (Thunb.) Makino (G. pentaphyllum) is an oriental herb documented to treat many diseases, including obesity, hyperlipidemia, metabolic syndromes and aging. However, the anti-obesity mechanism of G. pentaphyllum remains poorly understood.
OBJECTIVE: To reveal the anti-obesity mechanism of G. pentaphyllum Extract (GPE) in High-Fat Diet (HFD)-induced obese mice through untargeted metabolomics, Real-Time Quantitative PCR (RT-qPCR), and immunohistochemical experiments. Additionally, to tentatively identify the active constituents through LC-MS/MS and molecular docking approaches.
METHODS: GPE was prepared using ethanol reflux and purified by HP-20 macroporous resins. The components of GPE were identified by Liquid Chromatography- Mass Spectrometry (LC-MS) system. Forty-two C57BL/6 J mice were randomly and evenly divided into six groups, with seven mice in each group: the control group, obese model group, Beinaglutide group (positive control), and GPE low, medium, and high-dose groups (50 mg/kg, 100 mg/kg, and 200 mg/kg of 80% ethanol extract). Body weight, liver weight, blood glucose, blood lipids, and liver histopathological changes were assessed. Untargeted metabolomics was employed to characterize metabolic changes in obese mice after GPE treatment. The expression of genes related to differential metabolites was verified using Real-Time Quantitative PCR (RT-qPCR) and immunohistochemical experiments. The constituents with anti-obesity effects from GPE were tentatively identified through molecular docking approaches.
RESULTS: A total of 17 compounds were identified in GPE. GPE significantly lowered body weight, total cholesterol (TC) and low-density lipoprotein cholesterol (LDL-C) in obese mice and reduced liver weight and hepatic steatosis. Serum metabolomics identified 20 potential biomarkers associated with GPE treatment in obese mice, primarily related to tryptophan metabolism. GPE treatment downregulated the expression of Slc6a19 and Tph1 and upregulated Ucp1 expression. Molecular docking illustrated that compounds such as 20(R)-ginsenoside Rg3, Araliasaponin I, Damulin B, Gypenoside L, Oleifolioside B, and Tricin7-neohesperidoside identified in GPE exhibited favorable interaction with Tph1.
CONCLUSIONS: The extract of G. pentaphyllum can inhibit the absorption of tryptophan and its conversion to 5-HT through the Slc6a19/Tph1 pathway, upregulating the expression of Ucp1, thereby promoting thermogenesis in brown adipose tissue, facilitating weight loss, and mitigating symptoms of fatty liver. Triterpenoids such as Araliasaponin I, identified in GPE, could be the potential inhibitor of Tph1 and responsible for the anti-obesity activities.