基于~1H NMR技术的卵巢静止奶牛血浆代谢谱分析

发布时间：2018-06-26 13:37

本文选题：奶牛 + 乏情　；参考：《黑龙江八一农垦大学》2015年硕士论文

【摘要】：奶牛产后乏情是奶牛产后常发的一种繁殖障碍问题，是指奶牛分娩后胎衣排出体外开始到其生殖器官恢复正常的一段时间内（通常产后60-90天），泌乳奶牛没有明显的发情表现，并且卵巢无周期性机能活动状况。近年来，随着我国奶牛养殖规模不断增加，单产水平不断提高，奶牛产后乏情也日益增高，给奶牛生产带来了巨大的经济损失。尽管奶牛乏情与许多因素有关，包括营养、代谢和内分泌因素等，但是有关高产奶牛产后乏情与机体整体代谢的关系尚未明确。因此，，本试验应用代谢组学理论和技术——核磁共振技术结合多元统计分析和生物信息学分析，获得了卵巢静止所致的乏情奶牛血浆特征代谢物，明确了奶牛产后乏情与机体特征代谢物的关系，为进一步深入揭示奶牛产后乏情的代谢调节机制提供科学的理论依据。本试验在黑龙江省某集约化奶牛场随机选取产后60-90天的年龄、胎次、体况相近的健康经产高产奶牛（年产10t以上）共206头作为试验动物；根据奶牛临床表现、直检和B超检查结果以及激素检测结果选取10头奶牛为发情组（E），10头奶牛为乏情组（A）——卵巢静止；然后对两组奶牛血浆样品进行无偏性的一维氢谱核磁共振（1H Nuclearmagnetic resonance，1H NMR）检测，对比其血浆代谢组学图谱，进行主成分分析（Principlecomponent analysis, PCA）、偏最小二乘法-判别分析（Partial least squares-discriminant analysis,PLS-DA）和正交校正偏最小二乘法-判别分析（Orthogonal correction partial leastsquares-discriminant analysis, OPLS-DA）等多元统计分析，筛选及鉴定乏情奶牛血浆差异代谢产物，并进行生物信息学分析。结果显示：相对于正常发情奶牛，乏情奶牛血浆中共12种代谢产物表现异常，包括水平增加的乙酸、柠檬酸和酪氨酸，以及水平下降的低密度脂蛋白、极低密度脂蛋白、脂质、丙氨酸、丙酮酸、肌酸、胆碱、磷酸胆碱和甘油磷酸胆碱等；这些代谢产物与糖代谢、氨基酸代谢、脂蛋白代谢和胆碱代谢的异常密切相关；通过参与氰基氨基酸代谢通路、丙酮酸代谢（或糖酵解和糖异生）通路、甘氨酸，丝氨酸和苏氨酸代谢（或三羧酸循环）通路、三羧酸循环（TCA循环）通路、甘氨酸，丝氨酸和苏氨酸代谢通路、甘油磷脂代谢和甘氨酸，丝氨酸和苏氨酸代谢通路、甘油磷脂代谢通路、甘油磷脂代谢通路、泛醌的生物合成及其他萜类化合物和苯丙氨酸、酪氨酸和色氨酸的生物合成通路调节等，干扰了奶牛正常发情；同时，乏情奶牛往往处于能量负平衡状态，且可能伴发酮病和脂肪肝病；血浆肌酸、胆碱浓度降低和酪蛋白浓度升高是奶牛乏情的新变化。结论：应用1H NMR技术结合多元统计分析能够有效的筛选出患卵巢静止的乏情奶牛血浆差异代谢产物，全景式、动态性地阐明了奶牛产后乏情体内代谢紊乱状态，为奶牛产后乏情的深入研究奠定理论和方法学基础。
[Abstract]:Postpartum estrus in dairy cows is a common reproductive disorder after delivery. It refers to the period between the excrete of the fetal coat after delivery and the recovery of the reproductive organs (usually 60-90 days after delivery), and the lactating cows have no obvious estrus performance. And the ovary has no periodic functional activity. In recent years, with the continuous increase of the scale of dairy cattle breeding in China, the level of unit yield and the postpartum estrus of dairy cows are also increasing, which has brought huge economic losses to the production of cows. Although estrus is related to many factors, including nutrition, metabolism and endocrine factors, the relationship between postpartum estrus and body metabolism in high-yielding cows is not clear. Therefore, the metabolic theory and technique nuclear magnetic resonance (NMR) technique combined with multivariate statistical analysis and bioinformatics analysis were used to obtain the plasma characteristic metabolites of ovariectic cows caused by ovarian motility. The relationship between postpartum estrus and body characteristic metabolites in dairy cows was clarified, which provided a scientific theoretical basis for further revealing the metabolic regulation mechanism of postpartum estrus in dairy cows. A total of 206 healthy and high-yielding dairy cows (over 10 tons per year) were randomly selected from an intensive dairy farm in Heilongjiang Province for 60 to 90 days postpartum age, birth order and body condition, according to the clinical manifestations of the cows, a total of 206 cows with high yield (annual production of more than 10 tons) were selected as experimental animals. The results of direct examination, B-ultrasound examination and hormone test were 10 cows as estrus group (E) and 10 cows as oestrous group (A), then the plasma samples of both groups were detected by 1H nuclear magnetic resonance (1H NMR), and the plasma samples of the two groups were detected by 1H nuclear magnetic resonance (1H NMR). The plasma metabonomics maps were compared and analyzed by principal component analysis (PCA), partial least squares-discriminant analysis (PLS-DA) and orthogonal correction partial leastsquares-discriminant analysis (OPLS-DA). The plasma metabolites of estrous cows were screened and identified, and bioinformatics analysis was carried out. The results showed that the plasma levels of 12 metabolites were abnormal in estrous cows, including increased levels of acetic acid, citric acid and tyrosine, and decreased low density lipoprotein, very low density lipoprotein and lipid. Alanine, pyruvate, creatine, choline, choline phosphate and choline glycerophosphate; these metabolites are closely related to abnormal metabolism of glucose, amino acid, lipoprotein and choline; by participating in the metabolic pathway of cyanoamino acids, Pyruvate metabolism (or glycolysis and glycosylation) pathway, glycine, serine and threonine metabolism (or tricarboxylic acid cycle) pathway, tricarboxylic acid cycle (TCA cycle) pathway, glycine, serine and threonine metabolic pathway, Glycerol phospholipid metabolism and glycine, serine and threonine metabolism pathways, glycerol phospholipid metabolism pathway, ubiquinone biosynthesis and other terpenoids and phenylalanine, The regulation of biosynthesis pathway of tyrosine and tryptophan interferes with the normal estrus of dairy cows; at the same time, the estrous cows are often in a negative energy balance state, and may be accompanied by ketosis and fatty liver disease; plasma creatine, The decrease of choline concentration and the increase of casein concentration are the new changes of estrus in dairy cattle. Conclusion: 1H NMR technique combined with multivariate statistical analysis can effectively screen plasma differential metabolites of cows with still ovarium. The metabolic disorder in postpartum estrous body of dairy cows is elucidated in a panoramic and dynamic manner. To lay a theoretical and methodological foundation for the further study of postpartum estrus in dairy cows.
【学位授予单位】：黑龙江八一农垦大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：S858.23

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