[关键词]
[摘要]
目的 基于中药质量标志物(quality marker,Q-Marker)概念,在验证花椒温中止痛功效的基础上,分析花椒提取物在正常大鼠及模型大鼠的血中移行成分。方法 采用ig冰水结合冰浴方式建立寒邪犯胃型胃脘痛大鼠模型,并连续ig花椒提取物2周,观察大鼠一般情况,并对全血血细胞计数、脏器指数和胃组织病理变化进行检测。应用超高效液相色谱-四极杆-静电场轨道阱高分辨质谱对对照组、模型组、空白给药组、花椒高剂量组大鼠的血清样本数据进行采集,通过主成分分析和正交偏最小二乘判别法分析花椒温中止痛的潜在物质基础。通过PharmMapper反向对接筛选入血成分的作用靶点,并进行京都基因与基因组百科全书(Kyoto encyclopedia of genes and genomes,KEGG)通路富集分析;构建成分-靶点-通路网络图;应用分子对接技术对主要作用通路进行验证。结果 与模型组比较,花椒组大鼠耳廓颜色明显变红,全血白细胞、淋巴细胞及单核巨噬细胞计数明显升高(P<0.05),胸腺指数明显增加(P<0.01);胃组织局部坏死脱落和变性细胞减少。从空白给药组、花椒高剂量组大鼠血清中共鉴定出7个入血成分,其中4个为原型成分(羟基-α-山椒素、羟基-β-山椒素、羟基-ε-山椒素、二羟基-α-山椒素),3个为代谢产物;其中二羟基-α-山椒素仅存在于花椒高剂量组大鼠血清。花椒入血成分可以通过氢键等形式与过氧化物酶体增殖物激活受体(peroxisome proliferator activated receptor,PPAR)信号通路、Th17细胞分化相关的靶点蛋白[脂肪酸结合蛋白3(fatty acid binding protein 3,FABP3)、视黄醇类X受体β(retinoid X receptor beta,RXRB)、FABP7、Janus激酶3(Janus kinase 3,JAK3)]良好结合。结论 花椒可能是通过PPAR、Th17细胞分化信号通路调节免疫系统发挥温中止痛功效;羟基山椒素类化合物可作为花椒温中止痛功效的潜在Q-Marker进行深入研究。
[Key word]
[Abstract]
Objective Based on concept of quality marker (Q-Marker) of traditional Chinese medicine, middle-warming and pain- alleviating efficacy of Huajiao (Zanthoxyli Pericarpium) was verified, and then the migration components of its in blood of normal rats and model rats were analyzed. Methods Rat model of epigastric pain caused by cold pathogens invading the stomach was established by ig ice water combined with ice bath, and Zanthoxyli Pericarpium extract was ig for 2 weeks. Pathological changes were detected, including rats in general, whole blood count, organ index and gastric tissue. UPLC-Q-Extractive-Orbitrap MS was used to collect the data of serum samples in control group, model group, blank administration group, and Zanthoxyli Pericarpium high-dose group. Principal component and orthogonal partial least squares discriminant method were employed to analyze the potential effective compound of Zanthoxyli Pericarpium on effect of middle-warming and analgesic efficacy. Furthermore, protein target of component in blood was screened by reverse docking with PharmMapper, and Kyoto encyclopedia of genes and genomes (KEGG) pathway enrichment analysis was performed; Component-target-pathway network was constructed; Molecular docking was applied to verify the main pathway. Results Compared with model group, auricles color of rats in Zanthoxyli Pericarpium group was obviously reddened, and counts of leukocytes, lymphocytes and mononuclear macrophages in blood were significantly increased (P < 0.05), and thymus index was increased (P < 0.01). Local necrotic exfoliation and cellular denaturation of gastric tissue were reduced. Seven components were identified from serum of rats in blank administration group and Zanthoxyli Pericarpium high-dose group, among which four were prototype components (hydroxy-α-sanshool, hydroxy-β-sanshool, hydroxy-ε-sanshool, dihydroxy-α-sanshool), three of which were metabolites; Among them, dihydroxy-α-sanshool was only present in serum of rats in Zanthoxyli Pericarpium high-dose group. The blood components of Zanthoxyli Pericarpium could bind well to peroxisome proliferator activated receptor (PPAR) signaling pathway and Th17 cell differentiation-related target proteins of fatty acid binding protein 3 (FABP3), retinoid X receptor β (RXRB), FABP7, Janus kinase 3 (JAK3) through hydrogen bonds. Conclusion Zanthoxyli Pericarpium may regulate the immune system through PPAR and Th17 cell differentiation signaling pathways to exert its middle-warming and pain-alleviating effect. Hydroxy-sanshool components can be used as Q-Marker for the effect of Zanthoxyli Pericarpium for in-depth research.
[中图分类号]
R285.5
[基金项目]
四川省中医药管理局项目(2020HJZX001)