Project description:The rapidly increasing number of engineered nanoparticles (NPs), and products containing NPs, raises concerns for human exposure and safety. With this increasing, and ever changing, catalogue of NPs it is becoming more difficult to adequately assess the toxic potential of new materials in a timely fashion. It is therefore important to develop methods which can provide high-throughput screening of biological responses. The use of omics technologies, including metabolomics, can play a vital role in this process by providing relatively fast, comprehensive, and cost-effective assessment of cellular responses. These techniques thus provide the opportunity to identify specific toxicity pathways and to generate hypotheses on how to reduce or abolish toxicity.We have used untargeted metabolome analysis to determine differentially expressed metabolites in human lung epithelial cells (A549) exposed to copper oxide nanoparticles (CuO NPs). Toxicity hypotheses were then generated based on the affected pathways, and critically tested using more conventional biochemical and cellular assays. CuO NPs induced regulation of metabolites involved in oxidative stress, hypertonic stress, and apoptosis. The involvement of oxidative stress was clarified more easily than apoptosis, which involved control experiments to confirm specific metabolites that could be used as standard markers for apoptosis; based on this we tentatively propose methylnicotinamide as a generic metabolic marker for apoptosis.Our findings are well aligned with the current literature on CuO NP toxicity. We thus believe that untargeted metabolomics profiling is a suitable tool for NP toxicity screening and hypothesis generation.
Project description:MicroRNAs are important negative regulators of protein coding gene expression, and have been studied intensively over the last few years. To this purpose, different measurement platforms to determine their RNA abundance levels in biological samples have been developed. In this study, we have systematically compared 12 commercially available microRNA expression platforms by measuring an identical set of 20 standardized positive and negative control samples, including human universal reference RNA, human brain RNA and titrations thereof, human serum samples, and synthetic spikes from homologous microRNA family members. We developed novel quality metrics in order to objectively assess platform performance of very different technologies such as small RNA sequencing, RT-qPCR and (microarray) hybridization. We assessed reproducibility, sensitivity, quantitative performance, and specificity. The results indicate that each method has its strengths and weaknesses, which helps guiding informed selection of a quantitative microRNA gene expression platform in function of particular study goals.
Project description:The severe harm of depression to human life has attracted great attention to neurologists, but its pathogenesis is extremely complicated and has not yet been fully elaborated. Here, we provided a new strategy for revealing the specific pathways of abnormal brain glucose catabolism in depression, which from the supply of energy substrates and the evaluation of mitochondrial structure and function. By using stable isotope-resolved metabolomics technique, we discovered the tricarboxylic acid cycle (TCA cycle) is blocked and the gluconeogenesis is abnormally activated in chronic unpredictable mild stress (CUMS) rats. In addition, our results showed an interesting phenomenon that the brain attempted to activate all possible metabolic enzymes in energy-producing pathways, but CUMS rats still exhibited a low TCA cycle activity due to impaired mitochondria. Depression caused mitochondrial structure and function impaired, and then led to abnormal brain glucose catabolism. The combination of the stable isotope-resolved metabolomics and mitochondrial structure and function analysis can accurately clarify the mechanism of depression. The mitochondrial pyruvate carrier and acetyl-CoA maybe the key targets for depression treatment. The strategy provides a unique insight for exploring the mechanism of depression, the discovery of new targets, and the development of ideal novel antidepressants.
Project description:Oncogene-associated metabolic signatures in prostate cancer, identified by an integrative analysis of cultured cells and murine and human tumors, suggest that AKT activation results in a glycolytic phenotype whereas MYC induces aberrant lipid metabolism. Heterogeneity in human tumors makes this simplistic interpretation obtained from experimental models more challenging. Metabolic reprogramming as a function of distinct molecular aberrations has major diagnostic and therapeutic implications.
Project description:Prepartum exercise (PA) has been proposed as a strategy for the peripartum management of dairy cows; however, the mechanism by which PA affects metabolism has not been elucidated. Here, we investigated the metabolic changes in transition dairy cows with PA. Holstein transition multiparous dairy cows were assigned to an exercise (n = 12) or a control (n = 12) group; the cows in the exercise group walked for a targeted 45 min at 3.25 km/h, two times a day. Plasma non-esterified fatty acid (NEFA), β-hydroxybutyric acid (BHBA), glucose, and triglyceride levels were measured, and metabolic profiles were analyzed using untargeted mass spectrometry. Compared with those in the control group, the concentrations of NEFA at -7 d, glucose at 0 d, and BHBA at +7 d relative to calving were considerably decreased in the exercise group. Untargeted metabolomics analysis revealed differences in the levels of key metabolites, including kynurenine, tryptophan, homovanillic acid, dopamine, cis-9-palmitoleic acid, and palmitic acid, between the exercise and control group cows. This study suggests that PA may decrease homovanillic acid and cis-9-palmitoleic acid levels and increase tryptophan levels to alleviate the metabolic stress in dairy cows during calving, thereby improving postpartum health.
Project description:Cardiovascular diseases (CVDs) are noncommunicable diseases known for their complex etiology and high mortality rate. Oxidative stress (OS), a condition in which the release of free radical exceeds endogenous antioxidant capacity, is pivotal in CVC, such as myocardial infarction, ischemia/reperfusion, and heart failure. Due to the lack of information about the implications of OS on cardiovascular conditions, several methodologies have been applied to investigate the causes and consequences, and to find new ways of diagnosis and treatment as well. In the present study, cardiac dysfunction was evaluated by analyzing cells' alterations with untargeted metabolomics, after simulation of an oxidative stress condition using hydrogen peroxide (H2O2) in H9c2 myocytes. Optimizations of H2O2 concentration, cell exposure, and cell recovery times were performed through MTT assays. Intracellular metabolites were analyzed right after the oxidative stress (oxidative stress group) and after 48 h of cell recovery (recovery group) by ultra-high-performance liquid chromatography coupled to mass spectrometry (UHPLC-MS) in positive and negative ESI ionization mode. Significant alterations were found in pathways such as "alanine, aspartate and glutamate metabolism", "glycolysis", and "glutathione metabolism", mostly with increased metabolites (upregulated). Furthermore, our results indicated that the LC-MS method is effective for studying metabolism in cardiomyocytes and generated excellent fit (R2Y > 0.987) and predictability (Q2 > 0.84) values.
Project description:The rapid development of biogas plants in China has generated large quantities of digestate. The disparity between the continuity of biogas plant operation and the seasonality of digestate utilization has led to the need to store digestate. Therefore, untargeted profiling of bioactive compounds in the digestate stored under aerobic and anaerobic conditions was performed. The antioxidant and antifungal activity of digestate stored under varying conditions was likewise assessed. The results delineated that digestate storage under varying conditions brought about the degradation of organic acids, alkenes, aldehydes, alcohols, ketones, ethers, amino acids and their derivatives, and esters, leading to the stabilization of digestate components. Together, these new data revealed that digestate storage for up to 20 days under aerobic conditions promotes glycine, serine, and threonine degradation pathways and enhances biotin and vitamins production. In contrast, anaerobic storage enhances the taurine and hypotaurine metabolic pathways and increases the derivation of antimicrobial substances, such as indole alkaloids. Moreover, digestate storage under anaerobic conditions promotes antioxidant and antifungal activity more than storage under aerobic conditions. These findings can contribute to the future development of high-value agricultural products from digestate and the sustainability of biogas plants. Further studies are required for the untargeted metabolomic of digestate under storage to explore the underlying mechanisms of promoting disease resistance by the digestate upon land application.
Project description:Bottromycin A2 is a structurally unique ribosomally synthesized and post-translationally modified peptide (RiPP) that possesses potent antibacterial activity towards multidrug-resistant bacteria. The structural novelty of bottromycin stems from its unprecedented macrocyclic amidine and rare β-methylated amino acid residues. The N-terminus of a precursor peptide (BtmD) is converted into bottromycin A2 by tailoring enzymes encoded in the btm gene cluster. However, little was known about key transformations in this pathway, including the unprecedented macrocyclization. To understand the pathway in detail, an untargeted metabolomic approach that harnesses mass spectral networking was used to assess the metabolomes of a series of pathway mutants. This analysis has yielded key information on the function of a variety of previously uncharacterized biosynthetic enzymes, including a YcaO domain protein and a partner protein that together catalyze the macrocyclization.