山羊慢性酒糟中毒血液代谢物分析及乙醇中毒通路关键基因mRNA转录水平变化

Analysis of Blood Metabolites in Goat Chronic Distiller’s Grain Poisoning and Expression Changes of Key Genes in Ethanol Poisoning Pathway

本试验旨在揭示慢性酒糟中毒对山羊血液代谢物及其显著性代谢通路——乙醇中毒代谢通路的影响,筛选潜在性诊断生物标志物,为该病神经系统紊乱发生机制及早期诊断提供参考资料。试验选取4月龄健康大足黑山羊与波尔山羊的杂交后代15只,随机分为3组,65%白酒糟组、75%白酒糟组、基础日粮对照组。每隔15 d,进行肝肾、钙磷及蛋白质代谢指标检测,结合临床症状判断山羊慢性酒糟中毒建模成功。采集血样,利用超高效液相色谱—四极杆飞行时间质谱联用技术进行差异代谢物分析并筛选差异代谢物代谢富集通路,研究差异代谢通路中关键基因的表达变化。代谢组学结果表明,慢性酒糟中毒引起山羊体内多巴胺(DA)和2-呋喃酰甘氨酸(2-FG)上调,前列腺素I 2(PGI_(2))下调;突触小泡周期、多巴胺能突触、乙醇中毒、牛磺酸和亚牛磺酸代谢、组氨酸代谢、苯丙氨酸、酪氨酸和色氨酸生物合成、三羧酸循环、鞘脂信号通路、cAMP信号通路等代谢通路均差异显著(P<0.05)。其中,乙醇中毒代谢途径变化显著,且影响因子>0.2;乙醇中毒代谢通路中,75%白酒糟组TH、DAT表达量显著降低(P<0.05),DDC、MAOA表达量呈降低趋势(P>0.05)。综上所述,慢性酒糟中毒可引起山羊血液中有机酸类、氨基酸类、脂质与类脂分子类、胺类、醇类、脂肪酸类及类固醇类代谢物的显著性改变,DA、2-FG、PGI_(2)可作为慢性酒糟中毒病的潜在性诊断生物标志物。差异代谢物富集的共同通路为乙醇中毒代谢途径,途径中DAT、TH mRNA表达下降,使机体DA浓度升高,进而引起中毒山羊的神经功能紊乱。

This experiment aims at investigating the effects of chronic ethanol poisoning on goat blood metabolites and its important metabolic route,screen prospective diagnostic biomarkers,and comprehend some of the etiology of nervous system disorders of the disease and early detection.Fifteen healthy 4-month-old local goats were randomly assigned to three groups:65%distiller’s grains,75%distiller’s grains,basic diet control group.Three sets of blood samples were obtained every 15 days during the trial for the detection of liver and kidney,calcium,and protein metabolism indicators.The model of goat distiller’s grains chronic poisoning was deemed successful when combined with clinical symptoms.Collect blood samples and screen differential metabolites utilizing ultra-high performance liquid chromatography-quadrupole time-of-flight mass spectrometry,analyze metabolic enrichment pathways of differential metabolites,and evaluate the expression of key genes on the significant differential metabolic pathways.The metabolome results showed that chronic distiller’s grain poisoning caused up-regulation of dopamine(DA)and 2-Furoylglycine(2-FG)and down-regulation of prostaglandin I 2(PGI_(2))in goats;synaptic vesicle cycle,dopaminergic synapses,Metabolic pathways such as ethanol toxicity,taurine,and hypotaurine metabolism,histidine metabolism,phenylalanine,tyrosine and tryptophan biosynthesis,tricarboxylic acid cycle,sphingolipid signaling pathway,and cAMP signaling pathway were Significant difference.Among them,the metabolic pathways of ethanol poisoning changed significantly,and the impact factor was>0.2.In the metabolic pathway of alcohol intoxication,the expressions of TH and DAT in group II were significantly decreased(P<0.05);the expressions of DDC and MAOA showed a decreasing trend(P>0.05).In summary,chronic lees intoxication can cause significant alterations in organic acids,amino acids,lipids,lipid-like molecules,amines,alcohols,fatty acids,and steroids metabolites in the blood of goats,and DA,2-FG,and PGI_(2) can be used as potential diagnostic biomarkers for chronic lees intoxication disease.The common pathway of differential metabolite enrichment is the ethanol intoxication metabolic pathway,in which DAT and TH mRNA expression is decreased,resulting in elevated DA concentrations in the organism,which in turn causes neurological dysfunction in intoxicated goats.