不同品种大豆培养基对蝙蝠蛾拟青霉菌丝体代谢成分的影响

以不同品种大豆为培养基对蝙蝠蛾拟青霉代谢成分影响进行研究,筛选出最佳大豆品种,为蝙蝠蛾拟青霉标准化生产提供参考。首先对15个不同大豆品种培养基的主成分含量进行测定,同时对各品种大豆培养基培养的蝙蝠蛾拟青霉菌丝体生物量、菌丝体腺苷含量及腺苷总量进行测定。采用非靶向代谢组学方法对不同品种大豆培养基培养的蝙蝠蛾拟青霉菌丝体中的代谢成分进行研究。结果表明,15个不同品种大豆豆饼中的粗蛋白和粗脂肪含量以及对应培养基培养的蝙蝠蛾拟青霉菌丝体腺苷含量均有显著性差异(P<0.05),其中9号品种大豆培养基培养的菌丝体腺苷含量最高为(1 089.35±25.51) mg/kg, 5号的最低为(60.68±9.56) mg/kg, 5号、9号和10号培养的菌丝体之间存在显著性差异(P<0.05)。代谢组学研究表明,5号、9号和10号菌丝体代谢成分相对含量差异较大。其中9号菌丝体丙氨酸、谷氨酸、天冬氨酸等氨基酸,水苏糖、半乳糖等糖类,以及三羧酸循环产物2-氧代丁酸、柠檬酸等代谢物相对含量显著高于5号和10号菌丝体。KEGG富集分析表明,9号培养基通过促进天冬氨酸及谷氨酸代谢通路,可增加天冬氨酸和...     

Metabolomics as a diagnostic tool for hepatology: validation in a naturally occurring canine model

Human hepatopathies are a diagnostic challenge, with many distinct diseases having similar clinical signs and laboratory findings. Naturally occurring canine hepatic disease provides an excellent model for human diseases and similar diagnostic dilemmas exist; differentiating canine congenital portosystemic vascular anomalies (PVA) from acquired hepatopathies is difficult and traditionally requires invasive diagnostic procedures. The emerging post-genomic science of metabolomics is concerned with detecting global changes of populations of endogenous low molecular weight metabolites in biological samples and offers the possibility of identifying surrogate profiles of disease. Metabolomics couples sensitive metabolite analysis with sophisticated pattern recognition techniques. In this study, a metabolomic strategy has been employed to assess metabolite changes in the plasma of dogs with congenital PVA and acquired hepatic disease. Plasma samples were collected from 25 dogs, comprising 9 dogs with congenital PVA, 6 with acquired hepatopathy and 10 with non-hepatic disorders. Low molecular weight metabolites were analyzed by liquid chromatography-mass spectrometry (LC-MS). Following identification of metabolites, multivariate data analysis was used to compare profiles amongst groups. The analysis demonstrated significant disturbances in the plasma bile acid and phospholipid profiles of dogs with portovascular anomalies. In contrast to traditional laboratory parameters, the metabolomic strategy was able to produce a clear segregation between all three study groups. In conclusion, this study demonstrates the potential of metabolomics as a diagnostic tool for naturally occurring hepatic disease. With further validation, this approach will improve diagnostic capabilities, provide an insight into pathogenetic mechanisms, and ultimately inform therapeutic decision making in clinical hepatology. This work was supported in part by grants from The Royal Society, Petplan Charitable Trust and The Waltham Foundation.     

Combination of ‘omics’ data to investigate the mechanism(s) of hydrazine-induced hepatotoxicity in Rats and to identify potential biomarkers

To gain novel insight into the molecular mechanisms underlying hydrazine-induced hepatotoxicity, mRNAs, proteins and endogenous metabolites were identified that were altered in rats treated with hydrazine compared with untreated controls. These changes were resolved in a combined genomics, proteomics and metabonomics study. Sprague–Dawley rats were assigned to three treatment groups with 10 animals per group and given a single oral dose of vehicle, 30 or 90 mg?kg^?1 hydrazine, respectively. RNA was extracted from rat liver 48 h post-dosing and transcribed into cDNA. The abundance of mRNA was investigated on cDNA microarrays containing 699 rat-specific genes involved in toxic responses. In addition, proteins from rat liver samples (48 and 120/168 h post-dosing) were resolved by two-dimensional differential gel electrophoresis and proteins with changed expression levels after hydrazine treatment were identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry peptide mass fingerprinting. To elucidate how regulation was reflected in biochemical pathways, endogenous metabolites were measured in serum samples collected 48 h post-dosing by 600-MHz ¹H-NMR. In summary, a single dose of hydrazine caused gene, protein and metabolite changes, which can be related to glucose metabolism, lipid metabolism and oxidative stress. These findings support known effects of hydrazine toxicity and provide potential new biomarkers of hydrazine-induced toxicity.     

款冬花不同发育阶段的代谢组学和比较转录组学分析

以不同发育阶段款冬花（发育初期、中期、中后期、解封期及花朵期）为对象,基于核磁共振的代谢组学技术,分析其次生代谢物种类及含量的合成累积规律,并进行高通量转录组学测序,从差异表达的基因中寻找次生代谢物生物合成的关联酶基因。代谢组学分析发现,款冬花不同发育阶段的次生代谢物代谢组成明显不同,苯丙素类成分在发育初期至中后期含量较高,之后逐渐降低;黄酮类成分（芦丁、山奈酚）在发育的各个阶段含量均有波动,但总体变化不大。转录组学分析结果显示,与苯丙素类成分生物合成相关的酶基因（COMT、HCT）,随着花蕾的发育表达量逐渐降低;与黄酮类成分合成相关的酶基因（FLS、F3H、DFR）,其表达量在不同阶段变化不大。转录组测序中的相关酶基因表达量同次生代谢物成分的变化趋势基本一致。本文采用的代谢组学和转录组学结合的方式,对款冬花的次生代谢物累积规律进行分析,为今后款冬花次生代谢物的生物合成调控研究奠定了基础。     

兰茂牛肝菌子实体发育的关联物质及其主要通路分析

【背景】兰茂牛肝菌(Lanmaoaasiatica)等外生菌根真菌的子实体形成和发育机制仍然未知。【目的】揭示调控子实体发育的关联物质。【方法】同时运用核磁共振、气相质谱和液相质谱3种代谢组学技术,分析兰茂牛肝菌纯培养8d原基(Y8)与野生子实体(Z0)的小分子物质。【结果】Y8及Z0分别共指认出451、473种化合物;Y8vs.Z0,有362种显著或极显著上调(206种)及下调(156种)差异物质,其涉及47条调控通路。【结论】推测通过9条主要通路完成物质的深度转化及调控,极显著上调及下调差异物质如牛肝菌素可能对子实体的发育起着一定的调控作用,3种方法互相补充扩大了检测的泛度及灵敏度,这为探究兰茂牛肝菌子实体发育机理及人工培养提供了一定的理论参考。     

代谢组学技术在水产动物疾病研究中的应用

代谢组学是定量描述生物内源性代谢物质的整体及其对内因和外因变化应答规律的的一门新学科。近年来,代谢组学技术在水产动物疾病中的研究备受关注,特别是为感染性疾病发生机制及防控研究提供了一种新的手段。本文介绍了代谢组学技术及其在水产动物研究中的应用,包括代谢组学技术在水产动物感染性疾病、细菌耐药及环境应激等方面应用进行综述,分析了代谢组学在水产动物疾病研究中面临的问题与挑战,并对未来水产动物代谢组学研究趋势进行了展望,以期为代谢组学技术在水产动物疾病发病机制和药物研发方面更深入的运用提供参考。     

东亚钳蝎氯毒素通过下调PKM2介导的有氧糖酵解抑制神经胶质瘤细胞的增殖活性

东亚钳蝎氯离子通道毒素(Buthus martensii Karshch chlorion channel toxin,BmK CT)是一类短链神经毒素,在体内和体外均能有效抑制神经胶质瘤细胞侵袭和增殖。然而,BmK CT抑制胶质瘤细胞增殖活性的机制尚不清楚。本研究采用代谢组学的方法探索BmK CT抑制人脑神经胶质瘤细胞增殖的分子机制。首先,通过蛋白质表达纯化获得带有谷胱甘肽S转移酶(glutathione S-transferase, GST)标签的BmK CT蛋白和GST蛋白。利用圆二色谱检测两种蛋白质均具有α-螺旋二级结构,并且融合蛋白质GST-BmK CT热稳定性较好。通过噻唑蓝(methylthiazolyldiphenyl-tetrazolium bromide, MTT)法检测GST-BmK CT对神经胶质瘤细胞(U251和A172)增殖的影响。结果显示,与GST处理组相比,1μmol/L GST-BmK CT处理组对U251细胞的抑制率更明显(U251细胞19±1.1 vs. 2.25±0.95,P<0.001, 48 h; A172细胞12±1.4 vs. 2.25±1.25,P<0.001, 48 h)。利用气相色谱-质谱联用仪(gas chromatography-mass spectrometry, GC-MS)分析鉴定出细胞内的63种代谢物,通过通路分析和聚类分析证实,GST-BmK CT处理影响神经胶质瘤细胞的有氧糖酵解。根据峰面积统计数据显示,GST-BmK CT处理降低胞内丙酮酸的含量。GST-BmK CT处理组相比GST组,明显下调胞外培养基的乳酸分泌(16.33±3.78 vs. 35.6±2.31,P<0.001, 48 h),葡萄糖消耗(2.04±0.09 vs. 2.64±0.04,P<0.05, 48 h)和胞内腺苷三磷酸(ATP)含量(633.79±0.001 vs. 5392.71±266.67,P<0.05, 48 h)。结果表明,GST-BmK CT通过下调胶质瘤细胞有氧糖酵解水平降低ATP的生成。实时荧光定量PCR和蛋白质免疫印迹结果显示,GST-BmK CT降低U251细胞丙酮酸激酶(pyruvate kinase M2, PKM2)在转录水平和翻译水平的表达,进一步发现GST-BmK CT通过下调低氧诱导因子(hypoxia inducible factor-1α, HIF-1α)降低PKM2在转录水平的表达。以上结果表明,BmK CT通过下调HIF-1α的表达,降低PKM2介导的有氧糖酵解从而抑制神经胶质瘤细胞的增殖活性。     

Metabonomic models of human pancreatic cancer using 1D proton NMR spectra of lipids in plasma

In this study, we hypothesized that the altered insulin and glucose levels in male pancreatic cancer patients reported in a recent JAMA article would result in an altered lipid profile in the blood of pancreatic cancer patients when compared to controls (Stolzenberg-Solomon et al., 2005). Proton nuclear magnetic resonance (NMR) spectra of human lipophilic plasma extracts were used in order to build partial least squares discriminant function (PLS-DF) models that classified samples as belonging to the pancreatic control group or to the pancreatic cancer group. The sensitivity, specificity, and overall accuracy of the PLS-DF models based on 4 bins were 96%, 88%, and 92%, respectively. The sensitivity, specificity, and overall accuracy of the PLS-DF models based on 5 bins were 98%, 94%, and 96%, respectively. The sensitivity, specificity and overall accuracy of both the 4-bin and 5-bin PLS-DF models dropped only 1–2% during leave-25%-out cross-validation testing. Mass spectrometric profiling of phospholipids in plasma found three phosphatidylinositols that were significantly lower in pancreatic cancer patients than in healthy controls. The cancer models are based upon changes in lipid profiles that may provide a more sensitive and accurate diagnosis of pancreatic cancer than current methods that are based upon a single biomarker.     

NMR-based metabonomic analysis on effect of light on production of antioxidant phenolic compounds in submerged cultures of Inonotus obliquus

This study was designed to investigate the light effect on biosynthesis of antioxidant phenolic compounds by Inonotus obliquus grown in submerged cultures using ¹H NMR spectroscopy combining multivariate pattern recognition strategies. I. obliquus were exposed to a range of light conditions and resultant data were compared to those from field-grown sclerotia and the mycelia grown in daylight. Daylight illumination inhibited biosynthesis of davallialactone and phelligridins and other hispidin analogs. Continuous darkness enhanced the formation of phelligridins, davallialactone and inoscavins. Phelligridins and davallialactone also occurred in the mycelia grown in blue and red light with levels lower than those found in darkness. In addition, polyphenols synthesized under daylight conditions showed less potential antioxidant activity than those determined with other light regimes. These findings demonstrate that light regulates biosynthesis of polyphenols in I. obliquus and their subsequent antioxidant activities, and ¹H NMR-based metabolic profiling is a cost-effective approach for evaluating light effects on fungal metabolisms.     

附子化学成分和药理作用研究进展

长柱红山茶、美丽红山茶、贵州红山茶、皱叶瘤果茶和小黄花茶是颇具经济价值的稀有濒危植物,局部分布于贵州高原亚地区（ⅢD10 d）常绿阔叶林和常绿落叶阔叶混交林中。针对其物种濒危以及种子繁殖困难等问题,该研究对其种子生物学特性进行了分析。结果表明：（1）长柱红山茶千粒重最大（3 289.70 g）、发芽率最高（81.67%）,贵州红山茶千粒重最小（786.33 g）、发芽率最低（46.00%）;（2）小黄花茶含水率最高(48.85%),长柱红山茶含水率最低（39.52%）;（3）长柱红山茶生活力最大（98.33%）,皱叶瘤果茶生活力最小（63.33%）;（4）5种供试山茶种子的千粒重与生活力、发芽率、发芽势、发芽指数呈极显著正相关（R≥0.772,P＜0.01）,种子生活力与发芽率和发芽势呈极显著正相关（R≥0.738,P＜0.01）、与发芽指数呈显著正相关（R=0.532,P＜0.05）。以上结果说明,种皮较坚硬而不易吸水萌发,种子遭遇动物摄食和病虫侵蚀损耗而不利于种群繁衍是其物种濒危的重要原因。该研究结果为其植物资源的保护和利用提供了参考。