叶生小皮伞联合温针对偏头痛大鼠神经递质及ET-1蛋白水平的影响及其作用机制

目的 探讨叶生小皮伞联合温针对偏头痛大鼠神经递质及内皮素(Endothelin,ET)-1蛋白水平的影响及其作用机制。方法 用随机法将40只大鼠分为5组,每组8只,空白组Blank(BL)组,假手术组]、模型组Model(MO)组,建立偏头痛大鼠模型]、给药组Drug(DR)组,给予叶生小皮伞治疗]、温针组Warm needle(WN)组,给予温针治疗]、联合组Joint(JO)组,给予叶生小皮伞联合温针治疗]; 采用苏木精-伊红(Hematoxylin eosin,HE)染色观察脑组织神经细胞形态学变化; 采用酶联免疫吸附法(Enzyme linked immunosorbent assay,ELISA)检测大鼠血浆ET-1、降钙素基因相关肽(Calcitonin gene-related peptide,CGRP)水平,硝酸还原酶法测定一氧化氮(Nitric oxide,NO)水平; 用ELISA检测大鼠血清肿瘤坏死因子-α(Tumor necrosis factor-α,TNF-α)、白介素-6(Interleukin-6,IL-6)、中性粒细胞趋化因子-1(Cytokine-induced neutrophil chemoattractant-1,CINC-1)水平及脑组织中神经递质多巴胺(Dopamine,DA)、5-羟色胺(5-hydroxytryptamine,5-HT)水平; 免疫印迹(Western blot,WB)检测海马组织脑源性神经营养因子(Brain derived neurotrophic factor,BDNF)、酪氨酸激酶受体B(Tyrosine kinase receptor B,TrkB)蛋白水平。结果 BL组脑组织神经细胞染色质均匀分布在细胞核内,细胞未发生固缩、肿胀、坏死等病理现象; 与BL组比较,MO组神经元周围出现空泡,细胞萎缩; 与MO组比较,WN组、DR组、JO组神经细胞形态得到一定修复,且JO组神经细胞的形态修复较好,空泡减少。与BL组比较,MO组大鼠血浆NO,ET-1,CGRP水平均明显升高(P<0.05); 与MO组比较,WN组、DR组、JO组中NO,ET-1,CGRP水平均降低(P<0.05); 与WN组、DR组比较,JO组NO,ET-1,CGRP水平,且JO组降低的最明显(P<0.05)。与BL组比较,MO组TNF-α,IL-6,CINC-1水平均显著升高(P<0.05); 与MO组比较,WN组、DR组、JO组TNF-α,IL-6,CINC-1水平均显著降低; 与WN组、DR组比较,JO组各项水平均降低,且JO组降低最为显著(P<0.05)。与BL组比较,MO组5-HT,DA水平明显降低; 与MO组比较,WN组、DR组、JO组5-HT,DA水平均升高; 与WN组、DR组比较,JO组5-HT,DA水平明显升高。与BL组比较,MO组BDNF,TrkB表达水平均较高(P<0.05); 与MO组比较,WN组、DR组、JO组BDNF,TrkB表达水平均有不同程降低; 与DR组、WN组比较,JO组BDNF,TrkB表达水平显著降低(P<0.05)。结论 叶生小皮伞联合温针可以降低偏头痛大鼠ET-1蛋白水平,促进5-羟色胺等神经递质的表达,其机制可能与降低BDNF,TrkB表达水平有关。

The effect of Marasmiusepiphyllus combined with warm acupuncture on neurotransmitter and ET-1 protein in migraine rats

Objective To explore the effect of Marasmiusepiphyllus and warm acupuncture on neurotransmitters and ET-1 protein level in migraine rats. Methods Rat were randomly divided into 5 groups(8 rats in each group): blank group(Blank, BL group: sham operation group), model group(Model, MO group, migraine model), drug group(Drug, DR group received Phyllanthus vulgaris treatment), warm needle group(Warm Needle, WN group received warm needle treatment), combined group(Joint, JO group received Phyllanthus vulgaris combination Warm needle therapy). HE staining was used to observe the morphological changes of nerve cells in the brain tissue. ELISA was used to detect plasma active substances in rat blood vessels to detect ET-1, CGRP content. Nitrate reductase method was used to determine NO content;ELISA was used to detect rat serum TNF-α, IL-6, CINC-1 levels and the content of neurotransmitter DA and 5-HT in brain tissue. Western blot was used to detect the protein levels of BDNF and TrkB in the hippocampus. Results The chromatin of nerve cells in the brain tissue of the BL group was evenly distributed in the nucleus. The cells did not undergo pyknosis, swelling, necrosis and other pathological phenomena. Compared with the BL group, vacuoles and cell atrophy appeared around the neurons in the MO group. Compared with the MO group, the morphology of nerve cells in the WN group, the DR group and the JO group was reserved. The morphology of the nerve cells in the JO group was better repaired. The number of vacuoles were reduced. Compared with the BL group, the content of NO, ET-1, CGRP in the MO group in plasma was significantly increased(P<0.05).Compared with the MO group, the contents of NO, ET-1, and CGRP in the WN group, DR group, and JO group were decreased(P<0.05). Compared with the WN group and the DR group, the levels of NO, ET-1, and CGRP in the JO group were decreased(P<0.05). Compared with the BL group, the levels of TNF-α, IL-6, and CINC-1 in the MO group were significantly increased(P<0.05). Compared with the MO group, the levels of TNF-α, IL-6 in the WN, DR and JO groups were significantly higher. The level of CINC-1 was significantly reduced. Compared with the WN group and the DR group, the levels of TNF-α, IL-6 in JO group was decreased(P<0.05).Compared with the BL group, the 5-HT and DA levels of the MO group were significant decreased. Compared with the MO group, the level of 5-HT, DA were increased in WN group, DR group, and JO group. Compared with BL group, the expression of BDNF and TrkB were higher in MO group(P<0.05).Compared with the MO group, the expression levels of BDNF and TrkB in the WN group, the DR group, and the JO group all decreased. Compared with the DR group and the WN group, the levels of BDNF and TrkBin JO group decreased significantly(P<0.05). Conclusion The combination of Marasmiusepiphyllus and warm acupuncture reduces the level of ET-1 protein and promotes the expression of serotonin and other neurotransmitters. The mechanism may be related to the reduction of BDNF and TrkB signals.