背景:玄参[XS]已用于治疗甲状腺功能亢进。然而,其治疗甲状腺功能亢进和甲状腺激素引起的肝肾损伤的有效物质和药理机制尚未阐明。
目的:本研究基于网络药理学预测和实验验证,探讨XS治疗甲状腺功能亢进和甲状腺激素所致肝肾损伤的药理物质基础和潜在机制。
方法:基于使用超高效液相色谱串联四重外活性轨道阱高分辨率精确质谱(UPLC-QE-HRMS)鉴定的31种体内XS化合物,网络药理学方法用于机制预测。构建了系统网络来识别潜在的分子靶标,生物过程(BP),和信号通路。建立了组分-目标-途径网络。小鼠通过管饲法给予左甲状腺素钠30d,然后用不同剂量的含或不含丙基硫氧嘧啶(PTU)的XS提取物处理30d。肝脏,和肾脏样本使用酶联免疫吸附测定(ELISA)和蛋白质印迹进行分析。
结果:总共31个原型,60个I相代谢物,和23在口服XS提取物后在大鼠血浆中初步鉴定出II相代谢物。鉴定了31种体内化合物与疾病之间的96种潜在的共同靶标。京都基因和基因组百科全书(KEGG)分析显示,Bcl-2,BAD,JNK,p38和ERK1/2是首要目标。含或不含PTU的XS提取物具有以下作用:抑制左甲状腺素引起的T3/T4/fT3/fT4;血清中TSH水平增加;甲状腺结构的恢复;通过提高抗氧化酶过氧化氢酶(CAT)的活性来改善肝脏和肾脏的结构和功能,超氧化物歧化酶(SOD),和谷胱甘肽过氧化物酶(GSH-Px);激活抗凋亡蛋白Bcl-2;抑制凋亡蛋白p-BAD;下调炎症相关蛋白p-ERK1/2,p-JNK,和p-p38;抑制促炎细胞因子TNF-α的聚集,IL-1β,和IL-6,以及肝脏中的免疫细胞。
结论:XS可以通过抗氧化剂治疗甲状腺激素引起的甲状腺功能亢进和肝肾损伤,抗炎,和抗凋亡特性。此外,血清药物化学分析显示五种活性化合物,即4-甲基邻苯二酚,Sugiol,丁香酚,乙酰苯胺,和油酸,在体内具有不同的代谢途径,并显示出作为有效治疗剂的潜力。
BACKGROUND: Scrophulariae Radix (Xuanshen [XS]) has been used for several years to treat hyperthyroidism. However, its effective substances and pharmacological mechanisms in the treatment of hyperthyroidism and thyroid hormone-induced liver and kidney injuries have not yet been elucidated.
OBJECTIVE: This study aimed to explore the pharmacological material basis and potential mechanism of XS therapy for hyperthyroidism and thyroid hormone-induced liver and kidney injuries based on network pharmacology prediction and experimental validation.
METHODS: Based on 31 in vivo XS compounds identified using ultra-performance liquid chromatography tandem quadruple exactive orbitrap high-resolution accurate-mass spectrometry (UPLC-QE-HRMS), a network pharmacology approach was used for mechanism prediction. Systematic networks were constructed to identify the potential molecular targets, biological processes (BP), and signaling pathways. A component-target-pathway network was established. Mice were administered levothyroxine sodium through gavage for 30 d and then treated with different doses of XS extract with or without propylthiouracil (PTU) for 30 d. Blood, liver, and kidney samples were analyzed using an enzyme-linked immunosorbent assay (ELISA) and western blotting.
RESULTS: A total of 31 prototypes, 60 Phase I metabolites, and 23 Phase II metabolites were tentatively identified in the plasma of rats following the oral administration of XS extract. Ninety-six potential common targets between the 31 in vivo compounds and the diseases were identified. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis revealed that Bcl-2, BAD, JNK, p38, and ERK1/2 were the top targets. XS extract with or without PTU had the following effects: inhibition of T3/T4/fT3/fT4 caused by levothyroxine; increase of TSH levels in serum; restoration of thyroid structure; improvement of liver and kidney structure and function by elevating the activities of anti-oxidant enzymes catalase (CAT),superoxide dismutase (SOD), and glutathione peroxidase (GSH-Px); activation anti-apoptotic proteins Bcl-2; inhibition the apoptotic protein p-BAD; downregulation inflammation-related proteins p-ERK1/2, p-JNK, and p-p38; and inhibition of the aggregation of pro-inflammatory cytokines TNF-α, IL-1β, and IL-6, as well as immune cells in the liver.
CONCLUSIONS: XS can be used to treat hyperthyroidism and liver and kidney injuries caused by thyroid hormones through its anti-oxidant, anti-inflammatory, and anti-apoptotic properties. In addition, serum pharmacochemical analysis revealed that five active compounds, namely 4-methylcatechol, sugiol, eugenol, acetovanillone, and oleic acid, have diverse metabolic pathways in vivo and exhibit potential as effective therapeutic agents.