小儿支气管哮喘中医证候学生物标记物代谢组学研究

发布时间：2018-01-14 20:28

本文关键词：小儿支气管哮喘中医证候学生物标记物代谢组学研究　出处：《南京中医药大学》2017年硕士论文　论文类型：学位论文

【摘要】：支气管哮喘(Bronchial asthma)是威胁儿童健康最常见的慢性疾病。由于环境、生活方式改变等因素,我国小儿哮喘发病呈逐年上升趋势,造成了巨大的家庭负担。运用中医药防治小儿哮喘历史悠长,防治效果明显,有着完善的理法方药体系。目前儿童哮喘辨证多分为发作期、迁延期与缓解期三期,风寒束肺证、痰热阻肺证等八个证型。在临床工作中,哮喘证型的判断多依据医师的个人经验,尚缺乏辨别不同证型的客观化指标。证候是中医学中特有的名词,在整个理论框架内处于核心地位。加速中医证候规范化建设,实现证候的客观化、标准化,是中医学迈向科学化、走向世界的先决条件。本研究立足于中医证候客观化、标准化,以小儿支气管哮喘为切入点,采用代谢组学技术试图揭示小儿哮喘不同证候间的物质基础,是证候表征微观化的一次有益的探索。目的:分析小儿哮喘发作期痰热阻肺证、非痰热阻肺证,缓解期肺脾气虚证尿液样本代谢谱,探寻小儿支气管哮喘中医证候学标记物。方法:纳入符合诊断标准的小儿哮喘发作期痰热阻肺证24例、非痰热阻肺证20例,缓解期肺脾气虚证40例,同时纳入健康小儿29例设为正常组。采集各组小儿尿液标本进行气相色谱与质谱联用技术(Gas Chromatography-Mass Spectrometer/Mass Spectrometer,GC-MS/MS)检测,同时将痰热阻肺证、非痰热阻肺证组尿液标本进行液相色谱与质谱联用技术(Liquid Chromatography-Mass Spectrometer/Mass Spectrometer,LC-MS/MS)检测,将所得数据进行正交偏最小二乘法(Orthogonal partial least squares-discriminant analysis,OPLS-DA)判别分析,变量重要性投影值(Variable importance in the projection,VIP)、非参数检验以及XCMS Online分析平台筛选、确定组间差异性代谢物,同时利用MetaboAnalyst平台分析确定异常代谢通路。结果:各组间均能够良好区分,一系列差异性代谢物及代谢通路被确定。1.与正常组相比,哮喘发作期患儿尿中肌醇、尿酸、硬脂酸等14种物质的含量下降,氨基丙二酸含量升高;主要涉及丙酮酸代谢,赖氨酸降解、生物合成等5条代谢通路;2.与发作期非痰热阻肺证组相比,GC-MS/MS结果显示:痰热阻肺证患儿尿中草酸、L-苏氨酸、嘧啶等9种物质的含量下降,主要涉及戊糖磷酸途径,肌醇磷酸代谢4条等代谢通路。LC-MS/MS结果显示:痰热阻肺证患儿尿中蛋氨酸、琥珀酸、赖氨酸、肌酸、犬尿氨酸、环腺苷酸等多种有机酸、氨基酸,以及多种脂类、酮类化合物含量下降,仅羟基吲哚乙醛含量升高,主要涉及生物素代谢、色氨酸代谢等5条代谢通路。3.与肺脾气虚证组相比,痰热阻肺证患儿尿中氨基丙二酸、脯氨酸、苯丙氨酸、棕榈酸等8种物质含量升高,肌醇、尿酸、硬脂酸含量降低,主要涉及精氨酸和脯氨酸,甘氨酸、丝氨酸和苏氨酸等4条代谢通路。结论:小儿哮喘发作期与健康小儿尿液代谢物具有差异;支气管哮喘发作期不同证型、哮喘不同时期不同证型间存在代谢标记物与代谢通路基础,可能是区分中医不同证候的本质,通过进一步验证优化或可作为小儿哮喘中医证候判断生物学标记物。
[Abstract]:Bronchial asthma (Bronchial asthma) is the most common chronic disease threat to children's health. Because of the environment, lifestyle changes and other factors, China's children with asthma increased year by year, caused a huge burden on the family. The use of Chinese medicine prevention and treatment of asthma in children has a long history, and obvious effect, with perfect prescription system at present children with asthma syndrome and more divided into the stage of attack and remission, delayed phase three, cold beam lung, syndrome of phlegm heat in the lung and other eight syndromes. In clinical work, on the basis of doctor's personal experience of asthma syndromes, lack of objective indicators to identify different types of syndromes. Is a noun characteristic in traditional Chinese medicine, is the core in the whole theoretical framework. To accelerate the construction of TCM standardization, to achieve the objective, the standardization of syndrome of traditional Chinese medicine, is a prerequisite to scientific, this study of the world. In TCM syndrome objective, standardized, in children with bronchial asthma as the breakthrough point, using metabonomics technology to reveal the material basis of different syndromes in children asthma, is a useful exploration of the characterization of Micro syndrome. Objective: analysis of phlegm heat obstructing lung syndrome in children asthma and non phlegm heat in the lung syndrome, alleviate lung temperdeficiency urine samples metabolic spectrum, explore the TCM syndrome of childhood bronchial asthma syndrome markers. Methods: patients met the diagnostic criteria of asthma in children 24 cases of phlegm heat in the lung syndrome onset, 20 cases of non phlegm heat in lung syndrome, lung and spleen qi deficiency syndrome remission in 40 cases, at the same time in healthy children 29 cases of normal group. Chromatography and mass spectrometry in urine specimens collected gas (Gas Chromatography-Mass Spectrometer/Mass Spectrometer, GC-MS/MS) detection, and the phlegm heat in lung syndrome, non phlegm heat in the lung syndrome group in urine specimens For liquid chromatography and mass spectrometry (Liquid Chromatography-Mass Spectrometer/Mass Spectrometer, LC-MS/MS) detection, the data of orthogonal partial least squares (Orthogonal partial least squares-discriminant analysis, OPLS-DA) discriminant analysis, variable importance projection value (Variable importance in the projection, VIP), non parametric test and XCMS Online analysis platform selection, determine the differences between groups of metabolites, and use the MetaboAnalyst platform to analyze and confirm the abnormal metabolic pathways. Results: both groups can good area, a series of specific metabolites and metabolic pathways were identified in.1. compared with the normal group, inositol, children with asthma in the urine uric acid, decreased content of stearic acid and other 14 kinds of substances, amino. Malonic acid content; mainly involved in pyruvate metabolism, lysine degradation, biological synthesis etc. 5 metabolic pathways; and 2. For non phlegm heat in the lung syndrome group compared to GC-MS/MS results showed that oxalic acid, phlegm heat in lung syndrome in children with urinary L- decreased content of threonine, pyrimidine and other 9 kinds of substances, mainly involved in the pentose phosphate pathway, inositol phosphate metabolism and other metabolic pathways of 4.LC-MS/MS showed that methionine, phlegm heat in lung syndrome in children with urinary succinate lysine, creatine, kynurenine, camp and other organic acids, amino acids, and a variety of lipids, decreased ketone content increased only hydroxyindole acetaldehyde content, mainly related to biological Sudache, compared tryptophan metabolism of 5 metabolic pathways of.3. with lung and spleen qi deficiency group, phlegm heat in lung syndrome in children with urinary in malonic acid, proline, phenylalanine, increased the content of 8 compounds of palmitic acid and inositol, uric acid, reduce the content of stearic acid, mainly involving arginine and proline, glycine, serine and threonine 4 metabolic pathways. Conclusion: Children with asthma and healthy children with urinary metabolite differences; period of different syndromes of asthma, metabolic markers and metabolic pathways of different syndromes of asthma in different periods, may be essential to distinguish different syndromes of traditional Chinese medicine, further tested through optimization or can be used as asthma syndromes to judge biological markers.

【学位授予单位】：南京中医药大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R272

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