【摘要】 为了探究自拟中药复方五知散对免疫应激雏鸡生长性能与免疫功能的影响,试验基于网络药理学与分子对接技术初步探究五知散缓解鸡免疫应激的作用机制,通过TCMSP数据库筛选五知散中五味子、金银花、知母、甘草的活性成分,在TCMSP数据库获得相对应的靶点;通过在线软件Venny 2.1获取五知散和鸡免疫应激的交集靶点,预测五知散缓解鸡免疫应激的靶点;采用David数据库和微生信平台对五知散缓解鸡免疫应激相关靶点进行GO功能富集分析与KEGG通路分析;将蛋白互作分析网络图中度值排名前两位的靶点及其对应的活性成分通过Autodocktools 1.5. 7软件进行分子对接。将200只雏鸡随机分为5组,每组4个重复,每个重复10只。1日龄起,空白组、模型组饲喂基础日粮,五知散高、中、低剂量组分别在日粮中添加2%(20 g/kg)、1%(10 g/kg)、0.5%(5 g/kg)复方五知散,在应激I期(试验第21天)、应激Ⅱ期(试验第35天)检测免疫应激雏鸡生产性能指标、脏器免疫指数、血清生化指标、血清炎症因子。结果表明:五知散含槲皮素、木犀草素、山奈酚及甘草查尔酮A等114个活性成分,可作用于B细胞淋巴瘤2(Bcl-2)、白细胞介素-6(IL-6)等75个靶点,影响病灶粘连、MAPK信号通路、细胞凋亡等相关通路,其核心靶点与主要活性成分的对接结合能均<-20.83 k J/mol,有较好的结合活性。与模型组相比,在试验22～28 d,五知散高、低剂量组平均日采食量(ADFI)显著提高(P<0.05);试验29～35 d,平均日增重(ADG)显著提高(P<0.05);试验第21天,五知散中、低剂量组血清内毒素(ET)活性显著或极显著降低(P<0.05或P<0.01),低剂量组血清皮质酮(CORT)浓度显著降低(P<0.05),血清Bcl-2浓度极显著降低(P<0.001);试验第35天,五知散高剂量组胸腺和法氏囊指数极显著升高(P<0.01),血清总蛋白(TP)浓度极显著升高(P<0.01),血糖(GLU)浓度显著升高(P<0.05),肿瘤坏死因子-α(TNF-α)、白细胞介素-1β(IL-1β)、IL-6及白细胞介素-8(IL-8)浓度显著或极显著降低(P<0.05或P<0.01),五知散中剂量组血清Bcl-2浓度极显著降低(P<0.01)。说明五知散可通过槲皮素、木犀草素等有效成分作用于Bcl-2、IL-6等靶点发挥缓解鸡免疫应激的作用,日粮中添加五知散可提高免疫应激下雏鸡的生长性能和免疫能力,且高剂量即添加2%复方五知散整体效果更好。更多还原

【Abstract】 In order to investigate the effects of Wuzhisan( WZS), a self-formulated traditional Chinese medicine compound, on the growth performance and immune function of immune-stressed chicks, the mechanism of WZS in relieving chickens’ immune stress was preliminary explored based on network pharmacology and molecular interworking technology in the experiment. The active ingredients of Schisandra chinensis, Lonicera japonica Thunb, Anemarrhenae Rhizom, and Glycyrrhiza glabra in WZS were screened through the TCMSP database, and the corresponding targets were obtained in the TCMSP platform. The intersection targets of WZS and chicken immune stress were predicted by the online software Venny 2. 1, and the targets of WZS alleviating chicken immune stress were predicted. GO functional enrichment analysis and KEGG pathway analysis were performed on the targets related to WZS alleviating chicken immune stress by the database of David and the platform of Microbiology. The targets at the top two rankings in the network diagram of the protein interactions and their corresponding active ingredients were docked by Autodocktools1. 5. 7 software. Two hundred chicks were randomly divided into five groups with four replicates per group and 10 chicks per replicate. From 1 day of age, animals in the blank and model groups(high, medium and low dose groups of WZS)were fed the basal diets and the basal diet supplemented with 2% compound WZS(20 g/kg), 1% compound WZS(10 g/kg), and 0. 5%compound WZS(5 g/kg), respectively. At the stage of stress Ⅰ(day 21 of the experiment) and stress Ⅱ(day 35 of the experiment), the production performance indexes, organ immunity indexes, serum biochemical indexes, and serum inflammatory factors of the chicks under immune stress were detected. The results showed that WZS contained 114 active ingredients, including quercetin, luteolin, kaempferol and licochalcone A, which could act on 75 targets, including B-cell lymphoma 2( Bcl-2) and IL-6, and affect the focal adhesion, MAPK signaling pathway, apoptosis and other related pathways. The docking binding energies of its core targets and the main active ingredients were all<-20. 83 kJ/mol, which had a good binding activity. Compared with the model group, the average daily feed intake(ADFI) in WZS high-dose and low-dose groups was significantly increased from 22 to 28 days(P< 0. 05), and the average daily weight gain( ADG) was significantly increased from 29 to 35 d(P<0. 05). On the 21st day of the experiment, the activity of serum endotoxin(ET) in WZS medium and low dose groups was significantly decreased(P<0. 05 or P<0. 01); the concentration of serum corticosterane(CORT) in WZS low dose group was significantly decreased(P<0. 05); the serum Bcl-2 concentration in the WZS high-dose group was significantly decreased(P<0. 001). On the 35th day of the experiment, the thymus and bursa indices in WZS high dose groups were significantly increased(P<0. 01);the concentration of serum total protein( TP,P< 0. 01) and blood glucose( GLU) was significantly increased(P< 0. 05), and the concentrations of tumor necrosis factor-α(TNF-ɑ), interleukin-1β( IL-1β), interleukin-6( IL-6) and interleukin-8( IL-8) were significantly or highly significantly reduced(P<0. 05 or P<0. 01); the serum Bcl-2 concentration in the medium-dose group of WZS was significantly decreased(P<0. 01). The results indicated that WZS could alleviate immune stress of chickens through quercetin, lignans and other active ingredients acting on Bcl-2, IL-6 and other targets, and the addition of WZS to diets could improve the growth performance and immune ability of chicks under immune stress, and the overall effect of adding 2% compound WZS was the best.更多还原