基于代谢组学的不明原因男性不育研究

发布时间：2018-04-01 09:36

本文选题：代谢组学　切入点：毒理　出处：《南京医科大学》2015年博士论文

【摘要】：第一部分代谢组学研究平台建设代谢组学是系统生物学中的重要工具,能够有效反应生物体当前的生存状态。代谢组信息的获得主要依靠各种强有力的高通量分析平台进行。本部分研究利用260种代谢物标准品,使用UPLC-Orbitrap超高效液相联用静电场轨道阱质谱,分别建立了基于反相C18和亲水作用色谱的代谢组学分析平台,代谢物峰形对称,保留时间分布均匀,质谱仪分辨率高,可操作性强。能够进行血清、尿液和精浆样本的代谢组学分析。本部分还应用该平台对西酞普兰的抗抑郁机理进行了非靶向代谢组学研究。模型组与对照组之间获得了了良好的区分,并且总计26种代谢产物被认定是抑郁症发病过程中的潜在生物标志物,其中24种代谢物在经过药物治疗之后逐渐回归正常水平,有10种代谢物可以被认为是评价西酞普兰抗抑郁效果的关键标志物。从代谢组学角度阐述了西酞普兰能够调整由慢性温和不可预知应激所造成的包括色氨酸代谢、能量代谢以及神经递质合成在内的代谢紊乱。本部分还使用GC-MS建立了分析游离脂肪酸组的非靶向代谢组学分析平台,能够使用25微升血样,分析全血中的35种游离脂肪酸,该方法简便,快速、稳定,适用于血中游离脂肪酸组的测定,并使用该方法进行了二硫化碳致心血管疾病的毒理作用研究。第二部分基于代谢组学的不明原因男性不育研究为了对不明原因男性不育的诊断和病因进行研究和探讨,我们对80例不明原因男性不育患者和80例对照的精浆进行了代谢组分析。我们利用代谢组学数据建立了OPLS-DA诊断模型,并且使用50例病例和50例对照的独立人群对所建立的模型进行了验证。结果表明,该诊断模型在验证人群中的灵敏度为82%,特异性为92%。通过比较UMI和对照组的差异代谢物,以及研究代谢物与精液质量参数的相关性,我们发现了4-羟基苯乙酸对不明原因男性不育有保护作用,且呈明显的剂量反应关系,同时该物质与精子的密度高度相关,是不明原因男性不育的重要标志物。通过代谢通路分析,我们发现不明原因男性不育患者的氨基酸代谢明显改变,表现为氨的生成量增加,同时精浆内氨基酸水平下降,从而影响到氨基酸-tRNA合成通路导致男性生育能力发生变化,同时多种氨基酸与精子密度呈负相关。
[Abstract]:The first part of the study platform of metabonomics is an important tool in systems biology. Can effectively reflect the current living state of organisms. The acquisition of metabolite information mainly depends on a variety of powerful high-throughput analysis platforms. By using UPLC-Orbitrap ultra-high performance liquid phase coupled with electrostatic field orbital trap mass spectrometry, a metabonomics analysis platform based on reversed-phase C18 and hydrophilic interaction chromatography was established respectively. The metabolites have symmetrical peak shape, uniform retention time distribution and high mass spectrometer resolution. Strong maneuverability. Can carry on the serum, This part also used the platform to study the antidepressant mechanism of citalopram. A good distinction was obtained between the model group and the control group. A total of 26 metabolites were identified as potential biomarkers during the onset of depression, 24 of which gradually returned to normal levels after drug therapy. Ten metabolites can be considered as key markers for evaluating the antidepressant effect of citalopram. From a metabonomics point of view, citalopram is able to regulate tryptophan metabolism, which is caused by chronic mild and unpredictable stress. Energy metabolism and neurotransmitter biosynthesis. This part also uses GC-MS to establish a non-target metabonomics analysis platform for the analysis of free fatty acids, which can use 25 microliter blood samples to analyze 35 kinds of free fatty acids in the whole blood. The method is simple, rapid, stable and suitable for the determination of free fatty acids in blood. The toxicological effects of carbon disulfide on cardiovascular diseases were studied using this method. The second part was based on the study of unknown cause male sterility based on metabonomics in order to study the diagnosis and etiology of unknown male sterility. We analyzed the seminal plasma of 80 unexplained male infertility patients and 80 controls. We established a OPLS-DA diagnostic model using metabonomics data. The results showed that the sensitivity and specificity of the diagnostic model were 82 and 92 respectively. The difference metabolites between UMI and control group were compared. And to study the correlation between metabolites and semen quality parameters, we found that 4-hydroxyphenylacetic acid has a protective effect on male infertility of unknown cause and has a significant dose-response relationship, and it is highly correlated with sperm density. Through the analysis of metabolic pathway, we found that the amino acid metabolism of male infertility patients with unknown cause was obviously changed, which was manifested by the increase of ammonia production and the decrease of amino acid level in seminal plasma. Therefore, the effect of amino acid-tRNA synthesis pathway on male fertility changes, and many amino acids have negative correlation with sperm density.
【学位授予单位】：南京医科大学
【学位级别】：博士
【学位授予年份】：2015
【分类号】：R698.2

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