Analysis of the metabolic footprint and tissue metabolome of placental villous explants cultured at different oxygen tensions reveals novel redox biomarkers

Pre-eclampsia (PE) is a multi-system disorder of pregnancy hypothesised to arise from circulating factors derived from an unhealthy placenta. Some changes in placental phenotype seen in PE can be reproduced by culture in altered oxygen (O(2)) tension. Currently, these circulating factors are unidentified, partly due to the complexity of maternal plasma. Investigation of factors released from placental tissue provides a potential method to identify bioactive compounds. Experimental strategies to study compounds present in a biological system have expanded greatly in recent years. Metabolomics can detect and identify endogenous and secreted metabolites. We aimed to determine whether metabolites could be identified in placental cultures with acceptable experimental variability and to determine whether altered O(2) tension affects the composition of the placental metabolome. In this study we used gas-chromatography-mass spectroscopy to determine the presence of metabolites in conditioned culture medium (CCM) and tissue lysates of placental villous explants cultured in 1, 6 and 20% atmospheric O(2) for 96h. This experimental strategy had an intra-assay variation of 6.1-11.6%. Intra and inter-placental variability were 15.7-35.8% and 44.8-46.2% respectively. Metabolic differences were identified between samples cultured in 1, 6 and 20% O(2) in both CCM and tissue lysate. Differentially expressed metabolites included: 2-deoxyribose, threitol or erythritol and hexadecanoic acid. We conclude that metabolomic strategies offer a novel approach to investigate placental function. When conducted under carefully controlled conditions, with appropriate statistical analysis, metabolic differences can be identified in placental explants in response to altered O(2) tension. Metabolomics could be used to identify changes in conditions associated with placental pathology.     

Analytical approaches to metabolomics and applications to systems biology

Phenotypic expression of renal diseases encompasses a complex interaction between genetic, environmental, and local tissue factors. The level of complexity requires integrated understanding of perturbations in the network of genes, proteins, and metabolites. Metabolomics attempts to systematically identify and quantitate metabolites from biological samples. The small molecules represent the end result of complexity of biological processes in a given cell, tissue, or organ, and thus form attractive candidates to understand disease phenotypes. Metabolites represent a diverse group of low-molecular-weight structures including lipids, amino acids, peptides, nucleic acids, and organic acids, which makes comprehensive analysis a difficult analytical challenge. The recent rapid development of a variety of analytical platforms based on mass spectrometry and nuclear magnetic resonance have enabled separation, characterization, detection, and quantification of such chemically diverse structures. Continued development of bioinformatics and analytical strategies will accelerate widespread use and integration of metabolomics into systems biology. Here, we will discuss analytical and bioinformatic techniques and highlight recent studies that use metabolomics in understanding pathophysiology of disease processes.     

Metabolic changes in rats after intragastric administration of MGCD0103 (Mocetinostat), a HDAC class I inhibitor

MGCD0103, an isotype-selective HDACi, has been clinically evaluated for the treatment of hematologic malignancies and advanced solid tumors, alone and in combination with standard-of-care agents. In this study, we developed a serum metabolomic method based on gas chromatography-mass spectrometry (GC-MS) to evaluate the effect of intragastric administration of MGCD0103 on rats. The MGCD0103 group rats were given 20, 40, 80 mg/kg of MGCD0103 by intragastric administration each day for 7 days. Pattern recognition analysis, including both principal component analysis (PCA) and partial least squares-discriminate analysis (PLS-DA) revealed that intragastric administration of MGCD0103 induced metabolic perturbations. As compared to the control group, the levels of L-alanine, L-isoleucine, and L-leucine of MGCD0103 group decreased. The results indicate that metabolomic methods based on GC-MS may be useful to elucidate side effect of MGCD0103 through the exploration of biomarkers (L-alanine, L-isoleucine, and L-leucine). According to the pathological changes of liver at difference dosage, MGCD0103 is hepatotoxic and its toxity is dose-dependent.     

基于代谢组学的胃癌组织和血浆的糖代谢研究

背景：肿瘤细胞糖酵解明显增强,但糖酵解对机体全身代谢的影响尚不明确。目的：研究胃癌患者组织和血浆的糖代谢中间产物变化．以建立胃癌局部代谢与机体全身代谢的联系。方法：收集20份慢性胃炎组织、17份胃癌组织、15份慢性胃炎血浆、15份胃癌血浆和15份胃癌术后血浆样本,应用气相色谱-飞行时间质谱（GC-TOFMS）方法检测糖代谢中间产物的变化。结果：共鉴定出13种糖代谢中间产物。与慢性胃炎组织相比,胃癌组织中麦芽糖、葡萄糖、甲基B-D-吡喃葡萄糖苷、6-磷酸果糖、核糖浓度明显降低而乳糖、乳酸、柠檬酸浓度明显升高（P〈0．05）。与慢性胃炎血浆相比,胃癌血浆葡萄糖、乳酸、琥珀酸、延胡索酸和苹果酸浓度明显降低而蔗糖、甲基β—D-吡喃葡萄糖苷浓度明显升高（P〈0．05）。与胃癌血浆相比,胃癌术后血浆蔗糖、半乳糖酸内酯、甲基β—D-吡喃葡萄糖苷浓度明显降低而葡萄糖、乳酸、柠檬酸、琥珀酸、延胡索酸、苹果酸明显升高（P〈0．05）。结论：胃癌组织存在糖酵解、三羧酸循环代谢异常,并对全身代谢产生了影响。基于GC-TOFMS的代谢组学是研究胃癌代谢轮廓的合适平台。     

Thinking on functional mechanism of acupuncture for inflammatory bowel diseases based on Metabolomics

Acupuncture has a good therapeutic effect in treatment of inflammatory bowel diseases （IBD）, but its functional mechanism has not been systematically explained. Metabolomics is the scientific study of dynamic chemical processes involving metabolites as well as metabolic response of living organisms. Metabolomics, a research method with integrity and dynamics, corresponds to the overall regulatory effect of acupuncture and is in line with the overall concept of traditional Chinese medicine （TCM） and the concept of homeostasis. In the recent years, metabolomics has been extensively applied to the clinical and experimental study of IBD, and its potential applied value has been unanimously acknowledged by the researchers. In this article, the application status of metabolomics in acupuncture is summarized, and the research ideas to study the mechanism of acupuncture in the regulation and control of IBD by metabolomics are preliminarily explored.     

逍遥散对肝郁脾虚证和肝损伤模型大鼠干预作用的代谢组学研究

目的：利用代谢组学方法探讨肝郁脾虚证和肝损伤模型的共同本质及逍遥散的干预作用。方法将40只SD大鼠按随机数字表法分为空白对照组、肝郁脾虚证模型组、肝郁脾虚证给药组、肝损伤模型组、肝损伤给药组,每组8只。采用夹尾激怒致肝郁联合番泻叶致脾虚的方法制备肝郁脾虚证模型,采用四氯化碳致肝损伤法制备肝损伤大鼠模型。肝郁脾虚证给药组、肝损伤给药组灌胃2 g/ml逍遥散液,其余各组灌胃等体积生理盐水,连续给药28 d。末次给药后12 h,采用气相色谱-质谱法分析各组大鼠血清代谢物谱及含量变化,采用 HE 染色法观察各组大鼠肝组织病理学变化。结果肝郁脾虚证模型组和肝损伤模型组大鼠肝组织均出现细胞核萎缩、崩解、变性坏死并伴有淋巴细胞浸润等病理改变,且血清中甘氨酸、琥珀酸、丙酸、蛋氨酸、谷氨酰胺、苯丙氨酸含量较空白对照组升高(P＜0.05)。逍遥散可降低血清中甘氨酸、琥珀酸、丙酸、蛋氨酸、谷氨酰胺、苯丙氨酸含量(P＜0.05)。结论逍遥散可调节一碳单位、含硫氨基酸、芳香族氨基酸等特殊代谢及三羧酸循环,发挥肝保护作用。     

Metabonomics study of the effects of pretreatment with glycyrrhetinic acid on mesaconitine-induced toxicity in rats

Ethnopharmacological relevance

Aconitum carmichaelii Debx. (Fuzi), a commonly use traditional Chinese medicine (TCM), has often been used in combination with Rhizoma Glycyrrhizae (Gancao) to reduce its toxicity due to diester diterpenoid alkaloids aconitine, mesaconitine, and hypaconitine. However, the mechanism of detoxication is still unclear. Glycyrrhetinic acid (GA) is the metabolite of glycyrrhizinic acid (GL), the major component of Gancao. In present study, the effect of GA on the changes of metabolic profiles induced by mesaconitine was investigated using NMR-based metabolomic approaches.

Materials and methods

Fifteen male Wistar rats were divided into a control group, a group administered mesaconitine alone, and a group administered mesaconitine with one pretreatment with GA. Their urine samples were used for NMR spectroscopic metabolic profiling. Statistical analyses such as orthogonal projections to latent structures-discriminant analysis (OPLS-DA), t-test, hierarchical cluster, and pathway analysis were used to detect the effects of pretreatment with GA on mesaconitine-induced toxicity.

Results

The OPLS-DA score plots showed the metabolic profiles of GA-pretreated rats apparently approach to those of normal rats compared to mesaconitine-induced rats. From the t-test and boxplot results, the concentrations of leucine/isoleucine, lactate, acetate, succinate, trimethylamine (TMA), dimethylglycine (DMG), 2-oxo-glutarate, creatinine/creatine, glycine, hippurate, tyrosine and benzoate were significantly changed in metabolic profiles of mesaconitine-induced rats. The disturbed metabolic pathways include amino acid biosynthesis and metabolism.

Conclusions

GA-pretreatment can mitigate the metabolic changes caused by mesaconitine-treatment on rats, indicating that prophylaxis with GA could reduce the toxicity of mesaconitine at the metabolic level.