基于粪便代谢组学的逍遥散抗抑郁作用机制

目的应用核磁共振氢谱(1H-NMR)代谢组学技术,研究慢性温和不可预知应激(CUMS)抑郁大鼠粪便中内源性代谢物及代谢通路的变化,并评价逍遥散的改善作用,探讨逍遥散抗抑郁作用机制。方法采用CUMS造模程序建立抑郁模型,采用1H-NMR技术结合多变量数据分析方法,研究CUMS抑郁大鼠粪便代谢物谱的变化、鉴定相关代谢标志物并构建代谢通路。结果共筛选10种与抑郁相关的潜在生物标志物;与对照组比较,模型大鼠粪便中天冬酰胺、天冬氨酸、乳酸和丙酸水平显著升高(P<0.05,P<0.01),而苯丙氨酸、酪氨酸、谷氨酸、谷氨酰胺、丙氨酸和脯氨酸水平显著降低(P<0.05,P<0.01)。与模型组相比,逍遥散能显著升高大鼠粪便中苯丙氨酸、酪氨酸、谷氨酸、谷氨酰胺和脯氨酸的水平,能显著降低天冬酰胺、乳酸和丙酸的水平。与对照组比较,CUMS造模后6条代谢通路发生显著变化,包括(1)氨基酰tRNA生物合成,(2)丙氨酸、天冬氨酸和谷氨酸代谢,(3)精氨酸和脯氨酸代谢,(4)谷氨酸和谷氨酰胺代谢,(5)苯丙氨酸代谢以及(6)丙酮酸代谢。逍遥散显著回调(2),(3),(5)和(6)代谢通路。结论逍遥散可通过调控氨基酸代谢、糖代谢和肠道微生物代谢等途径综合发挥抗抑郁作用,为全面、深入揭示逍遥散抗抑郁作用机制提供实验依据。     

基于核磁共振的代谢组学研究进展

基于核磁共振技术的代谢组学研究，是近几年发展起来的一种新的“组学”技术。它主要是利用生物体液的核磁共振谱图所提供的生物体内全部小分子代谢物的丰富信息，通过对这些信息的多元统计分析和模式识别处理，了解相关生物体在功能基因组学、病理生理学、药理毒理学等方面的状况及动态变化，以及它们所揭示的生物学意义，并从分子水平来认识生命运动的规律。本文综述了核磁共振技术在代谢组学方面的研究进展。     

基于核磁共振的代谢组学研究及其应用

基于核磁共振技术的代谢组学研究,是近几年发展起来的一种新的"组学"技术。本文介绍了代谢组学的定义、研究范畴和技术平台,着重阐述了基于核磁共振的代谢组学研究的基本原理、特点及其应用。     

基于~1H NMR血清和肝脏代谢组学的黄芩叶抗衰老作用研究

本实验采用D-半乳糖致衰老大鼠模型结合1H NMR血清和肝脏代谢组学,探究黄芩叶醇提物的抗衰老作用以及潜在抗衰老代谢调节机制。所有动物实验均通过山西大学科学研究伦理审查委员会审查。结果表明,黄芩叶醇提物具有抗衰老作用,能够改善衰老大鼠外在表征、自主活动能力和脂质过氧化、糖基化等衰老损伤,并可通过调节血清中谷氨酸、谷氨酰胺等12种差异代谢物和D-谷氨酰胺和D-谷氨酸代谢,丙氨酸、天冬氨酸和谷氨酸代谢等11个代谢通路,调节肝脏中α-葡萄糖、β-葡萄糖等5种差异代谢物和糖酵解或糖异生、淀粉和蔗糖代谢通路,改善衰老大鼠血清和肝脏代谢异常。     

NMR法研究小鼠乳腺癌生物标记物的代谢组学

目的用NMR法研究小鼠乳腺癌(Balb-c/EMT-6)不同生长周期中血清的代谢组学,寻找可能与肿瘤生长代谢相关的生物标记物。方法采集小鼠血清中代谢组分的NMR谱,对比小鼠乳腺癌肿瘤生长组(1～4周)与正常对照组代谢成分的变化,经绿原酸(CHA)治疗两周组的代谢物变化验证,结合PLS-DA模式识别分析,寻找小鼠乳腺癌肿瘤生长潜在的生物标志物。结果正常组与肿瘤生长组有明显的分离,肿瘤生长组中各周期组之间相互分离。两周CHA治疗组与正常组接近,但与肿瘤组明显分离。与正常组比较,肿瘤组的乳酸、乙酸和胆碱随肿瘤生长呈增加趋势,柠檬酸和肌酸的递增趋势较弱,而葡萄糖和脂蛋白(VLDL/LDL)随肿瘤生长有所减小。两周CHA治疗组与同期的肿瘤组比较,代谢关联组分发生明显变化(乳酸、胆碱和乙酸的量有所下降),与正常组基本接近。结论通过PLS-DA法提取出的乳酸、乙酸、胆碱等代谢组分与肿瘤生长具有一定相关性,可作为乳腺癌潜在的生物标志物的研究对象。     

基于NMR的免疫抑制小鼠模型血清代谢组学研究

目的采用核磁共振氢谱(~1H-NMR)代谢组学技术分析免疫抑制小鼠血清代谢物变化特点。方法将20只KM小鼠随机分为正常组和模型组,模型组用环磷酰胺建立免疫抑制模型,对其血清进行1H-NMR检测,采用正交偏最小二乘判别法(OPLS-DA)对NMR图谱进行分析,并确定两组小鼠血清差异性代谢物。结果与正常组小鼠比较,模型组小鼠血清谷氨酰胺、糖蛋白、不饱和脂类、VLDL、LDL、丙酮含量减少,亮氨酸、丙氨酸、酪氨酸、组氨酸、乳酸、甘氨酸、肌酸、蛋氨酸、缬氨酸、β-羟丁酸和肉碱含量增多。结论免疫抑制小鼠体内存在代谢网络的失调,具体表现为脂肪酸β-氧化、糖酵解和蛋白质分解过程加剧,脂肪动员被抑制。     

基于~1H-NMR护肝片抗大鼠急性肝损伤的代谢组学研究

目的寻找与四氯化碳(carbon tetrachloride,CCl_4)致大鼠急性肝损伤相关的潜在代谢标志物,并评价护肝片干预肝损伤的药效。方法大鼠腹腔注射CCl4建立急性肝损伤模型,检测血清天门冬氨酸氨基转移酶(aspartate aminotransferase,AST)、丙氨酸氨基转移酶(alanine aminotransferase,ALT)、碱性磷酸酯酶(alkaline phosphatase,ALP)和乳酸脱氢酶(lactate dehydrogenase,LDH)水平,同时采用核磁共振氢谱(1H-NMR)技术结合多变量数据分析方法,研究CCl_4所致大鼠尿液和粪便代谢物谱的变化并鉴定相关代谢标志物,从整体代谢表型和代谢标志物角度评价护肝片疗效。结果护肝片能明显降低血清中AST、ALT、ALP、LDH水平,且能有效逆转CCl_4所致大鼠尿液和粪便代谢紊乱,并分别明显回调5种尿液代谢标志物(α-酮戊二酸、柠檬酸、肌酐、氧化三甲胺、马尿酸)和3种粪便代谢标志物(丁酸、葡萄糖、尿嘧啶)。结论护肝片可能通过调节三羧酸循环和肠道菌群代谢起到抗急性肝损伤的作用。     

脾气虚证模型大鼠尿液的~1H-NMR代谢组学研究

目的建立脾气虚证大鼠模型,分析脾气虚证的代谢通路,探究内源性代谢产物变化与脾气虚证的关系。方法采用苦寒泻下、劳倦过度和饥饱失常复合因素方法建立并评价脾气虚证大鼠模型,检测其血清中肌酸磷酸激酶活性;对大鼠尿液进行~1H-NMR检测,以变化倍数(fold change,FC)>1.2,结合独立样本t检验(P<0.05)的统计学方法筛选脾气虚证模型组和对照组的差异代谢物,进行代谢通路分析与富集分析。结果脾气虚证模型建立成功;模型组的肌酸磷酸激酶活性低于对照组(P<0.05);从主成分分析(PCA)结果得出模型组和对照组的代谢产物得到明显区分;筛选出33种差异代谢物,主要参与的代谢通路涉及能量代谢、氨基酸代谢、核苷酸代谢,同时对肠道菌群存在干扰。结论脾气虚主要扰乱了大鼠的能量代谢通路(三羧酸循环、糖酵解、脂质氧化),抑制了供能作用,导致大鼠出现疲乏和体质量增长抑制的症状。     

应用核磁共振代谢组学方法分析食管癌患者血清代谢物

目的 应用核磁共振代谢组学方法研究食管癌患者血清代谢物的特征及其变化.方法 对健康人血清(健康组,10例)和食管癌患者血清(食管癌组,20例)样本检测核磁共振氢谱,运用统计学分析研究食管癌患者血清代谢组的特征及其改变.结果 食管癌患者血清中脂、乳酸和丙酮酸的含量升高,葡萄糖、三甲胺和胆碱/磷酸胆碱的含量降低(P<0.05).结论 血清代谢物的变化可以区分健康人和食管癌患者.这种基于核磁共振氢谱的代谢组学方法可为食管癌的诊断提供可靠代谢依据.     

基于多维组学探究逍遥散低极性部位对CUMS模型大鼠的抗抑郁作用

研究逍遥散低极性部位对抑郁模型大鼠干预作用,从肠道菌群和代谢物角度探讨逍遥散低极性部位抗抑郁作用机制。所有动物实验均通过山西大学科学研究伦理审查委员会审查。采用慢性温和不可预知应激(CUMS)程序对大鼠进行造模,以逍遥散低极性部位及阳性药(文拉法辛)为干预药物,结合16S rRNA基因测序和LC-MS代谢组学分析方法,探究逍遥散低极性部位对CUMS大鼠盲肠内容物肠道菌群和代谢物的影响,并对肠道菌群与代谢物进行Pearson关联分析。结果显示,逍遥散低极性部位显著改善CUMS大鼠的抑郁样行为;回调CUMS大鼠海马脑源性神经营养因子(BDNF)水平。肠道微生物群分析显示:逍遥散低极性部位给药可以增加CUMS模型大鼠微生物群的多样性,显著回调CUMS模型大鼠肠道微生物中罗斯氏菌属(Rothia)、普雷沃氏菌属([Prevotella]),其主要与肠道炎症和短链脂肪酸的产生有关。代谢组学结果表明,盲肠内容物中鉴定出与抑郁症相关的20种生物标志物,逍遥散低极性部位干预后可回调17种,涉及的通路为亚油酸代谢、牛磺酸和亚牛磺酸代谢、初级胆汁酸生物合成、精氨酸和脯氨酸代谢。相关分析进一步表明,逍遥散低...     

代谢物组学的研究进展

代谢物组学是继基因组学和蛋白质组学发展起来的一门新的组学(-omics)技术,是研究药物毒理和基因功能的技术平台,通过分析生物的体液、组织中的内源性代谢产物谱的变化来研究整体的生物学状况和基因调节功能.代谢物组学所涉及的主要技术为核磁共振技术和模式识别分析.对代谢物组学研究的样品、核心技术、分析方法和应用等几方面作简要概述.     

Solution structure of the phosphorylated sites of ribosomal protein S6 by 1H NMR spectroscopy

An increase in the rate of protein synthesis is found to be accompanied by phosphorylation of the 40S ribosomal protein S6. Treatment of S6 by cyanogen bromide produced three fragments, and one of the fragments of S6, which is a C-terminal portion of S6 (M_r? 4000), contains all phosphorylation sites of S6. The C-terminal fragment of S6 contains seven serines. S6 kinase phosphorylates S6 specifically, i.e. five serines in the C-terminal of S6 are phosphorylated. The three-dimensional structure of S6 peptide was studied in 50% trifluoroethanol/50% H₂O solution by ¹H NMR with combined use of distance geometry and restrained molecular dynamics calculations. NMR results indicated that it takes an α-helix between Glu5 and Arg21 and a distorted helical structure for the following three residues, but no rigid structure was present from Ser25 through the C-terminus and for the N-terminal region (Lys1-Lys4). The specificity of the phosphorylation of the peptide is discussed from a structural aspect. © Munksgaard 1996.     

Echistatin: the refined structure of a disintegrin in solution by 1 H NMR and restrained molecular dynamics

The structure of the disintegrin echistatin has been determined by ¹H NMR, distance geometry calculations and restrained molecular dynamics simulations. The structure has been refined from the preliminary distance geometry calculations with the inclusion of additional ¹H NMR data and hydrogen bonds identified in early stages of the molecular dynamics calculations. The calculations reported here allow a distinction to be made between the two possible disulfide bridging patterns - echistatin is crosslinked as follows: Cys2-Cysll, Cys7-Cys32, Cys8-Cys37, Cys20-Cys39. The final set of structures gives an average pairwise root mean square distance of 0.100 nm (calculated over the backbone atoms of residues Ser4-Cys20 and Asp30-Pro40). The core of echistatin is a well defined though irregular structure, composed of a series of non-classical turns crosslinked by the disulfide bridges and stabilised by hydrogen bonds. The RGD sequence is located in a protruding loop whose stem is formed by two rigid, hydrogen-bonded strands (Thr18-Cys20, Asp30-Cys32). The RGD sequence is connected to this structure by short, flexible segments. High (but not unlimited) mobility is probably necessary for fast recognition and fitting to the integrin receptors. Sequence variability among the disintegrins is found in the segments flanking the RGD sequence, suggesting that these may be important in conferring specificity for the receptors.     

胚胎干细胞的代谢组学方法在毒理学中的应用进展

胚胎干细胞(ESC)是高度未分化的细胞,具备体外无限增殖和诱导分化成三个胚层细胞的特性,所以胚胎干细胞可以作为一种毒性评价的工具。代谢组学是近年来发展起来的对某一生物或细胞内源性的所有低分子量代谢产物进行定量和定性分析的一门新学科,它以生物体液,细胞提取物,细胞培养液和组织等为研究对象,研究手段主要是核磁共振和质谱。该文综述了胚胎干细胞和代谢组学相关的实验技术及其在毒理学中的应用现状,同时对基于胚胎干细胞的代谢组学在毒理学研究中的应用和发展趋势进行了探讨。     

Survey and qualification of internal standards for quantification by H NMR spectroscopy

In quantitative NMR (qNMR) selection of an appropriate internal standard proves to be crucial. In this study, 25 candidate compounds considered to be potent internal standards were investigated with respect to the ability of providing unique signal chemical shifts, purity, solubility, and ease of use. The ¹H chemical shift (δ) values, assignments, multiplicities and number of protons (for each signal), appropriateness (as to be used as internal standards) in four different deuterated solvents (D₂O, DMSO-d₆, CD₃OD, CDCl₃) were studied. Taking into account the properties of these 25 internal standards, the most versatile eight compounds (2,4,6-triiodophenol, 1,3,5-trichloro-2-nitrobenzene, 3,4,5-trichloropyridine, dimethyl terephthalate, 1,4-dinitrobenzene, 2,3,5-triiodobenzoic acid, maleic acid and fumaric acid) were qualified using both differential scanning calorimetry (DSC) and NMR spectroscopy employing highly pure acetanilide as the reference standard. The data from these two methods were compared as well as utilized in the quality assessment of the compounds as internal standards. Finally, the selected internal standards were tested and evaluated in a real case of quantitative NMR analysis of a paracetamol pharmaceutical product.     

高血压病肝阳上亢证患者血清样品的核磁共振谱代谢组学研究

目的采用1HNMR结合主成分分析（PCA）方法研究高血压病肝阳上亢证患者和健康志愿者的血液成分谱差异,寻找可能的生物标记物,揭示高血压病肝阳上亢的证候本质。方法根据临床高血压病肝阳上亢证的诊断标准,选择典型病例,以1HNMR分析高血压病肝阳上亢证患者和健康志愿者的血液成分,使用PCA方法进行模式识别,寻找标记物并做出相关代谢途径的可能解释。结果PCA方法处理高血压病肝阳上亢证患者和健康志愿者的血清1HNMR谱数据显示,两组1HNMR谱数据可以在得分图实现分类,与健康志愿者作比较,患者机体相关代谢发生显著变化,体内部分氨基酸代谢和葡萄糖代谢明显异于健康志愿者。结论1HNMR结合PCA模式识别的代谢组学方法具有研究复杂条件下机体病理生理变化的优势,为理解高血压病肝阳上亢证的证侯本质和诊断该类病证提供了科学依据。     

Structures of neuropeptide γ from goldfish and mammalian neuropeptide γ, as determined by 1H NMR spectroscopy

Abstract: Neuropeptide γ belongs to tachykinin families which have a common C‐terminal amino acid sequence (Phe‐X‐Leu‐Met‐NH₂) and which induce various biological responses including salivation, hypotension, and contraction of gastrointestinal, respiratory, and urinary smooth muscle. In the present study, we present the solution structures of neuropeptide γ (NPγ) from gold fish (G‐NPγ) and mammalian NPγ (M‐NPγ), as determined by nuclear magnetic resonance (NMR) spectroscopy in 50% trifluoroethanol (TFE)/water (1 : 1, v/v) solution and 200 mm sodium dodecyl sulfate (SDS) micelles. In aqueous TFE solution, G‐NPγ has a α‐helical conformation in the region of His¹²–Met²¹ and a short helix in the N‐terminal region, and has a β‐turn from Arg⁹ to Arg¹¹ in between. In aqueous TFE solution, M‐NPγ also has α‐helical conformations both in the C‐terminal region and the N‐terminal region and a β‐turn from His⁹ to Arg¹¹ in between. In SDS micelle, the structure of G‐NPγ contains a stable α‐helix from His¹² to Met²¹ and a β‐turn from Arg⁹ to Arg¹¹, while M‐NPγ has a short helix from Ser¹⁶ to Met²¹. The region from His¹² to Met²¹ corresponds to the amino acid sequence of neurokinin A. Neuropeptide γ may act as a precursor of neurokinin A and the post‐translational processing of this peptide involves the enzymatic attack of the basic β‐turn region from residue 9 to residue 11 in the middle. From our relaxation study, it could be suggested that in fish system G‐NPγ induces the biological actions corresponding to those of substance P in mammalian system. The structures of G‐NPγ and M‐NPγ contain α‐helical structures at the C‐terminus and this helix seems to promote the affinity for NK₁ and/or NK₂ receptor.     

肾脏肿瘤组织的高分辨魔角旋转核磁共振~1H谱研究

目的 应用高分辨魔角旋转核磁共振~1H谱研究肾脏肿瘤组织的代谢变化.方法 选取肾癌肿瘤组织和癌旁对比组织标本,分别作病理分析和高分辨魔角旋转核磁共振~1H谱检测.结果 通过对比发现乳酸、胆碱类物质(胆碱、磷酸胆碱和甘油磷酸胆碱)及肌醇在肾癌组织中明显升高.结论 肾癌组织的代谢特征明显不同于癌旁对比组织,这可以为肾癌的诊断提供分子水平上的代谢依据.