<Superscript>1</Superscript>H NMR-based metabolomics approach reveals metabolic alterations in response to dietary imbalances in <Emphasis Type="Italic">Megalobrama amblycephala</Emphasis>

Introduction

High-fat and high-carbohydrate diets cause a number of metabolic disorders in mammals. However, little is known about metabolomic changes caused by dietary imbalances in fish.

Objectives

The objective of this study was to assess the impacts of high-fat diet (HFD), high-carbohydrate diet (HCD) and high-fat-high-carbohydrate diet (HFHCD) on metabolites in a farmed cyprinid fish Megalobrama amblycephala.

Methods

We have employed the ¹H NMR-based metabolomic approach to measure the concentrations of metabolites in plasma and liver of four different diet groups: HFD, HCD, HFHCD and control. Multivariate statistical analyses were used to determine significantly changed metabolites between all group-pairs.

Results

All three test diets have affected metabolic profiles, phenotypes and clinical chemistry. High-fat diets (HFD, HFHCD) resulted in a higher average weight than HCD, but high-carbohydrate diets (HCD, HFHCD) caused signs of liver damage. HCD has resulted in elevated metabolites in energy pathways, leading to further disturbances in creatine pathway. Excess of carbohydrate and lipid metabolism products in the HFHCD group appears to have caused “congestion” of the TCA cycle, causing a significant decline in the numbers of amino acids entering the cycle, which in turn resulted in elevated levels of seven amino acids in this group. Gut microbiota metabolites (TMA) exhibited a strong positive correlation with the carbohydrate content and a negative correlation with the fat content in diets.

Conclusion

These results provide an important insight into the diet-affected metabolic disorders that often lead to financial losses in the aquaculture of Megalobrama amblycephala.

Graphical Abstract

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Betaine reduces β-amyloid-induced paralysis through activation of cystathionine-β-synthase in an Alzheimer model of <Emphasis Type="Italic">Caenorhabditis elegans</Emphasis>

Background

The neurodegenerative disorder Alzheimer’s disease is caused by the accumulation of toxic aggregates of β-amyloid in the human brain. On the one hand, hyperhomocysteinemia has been shown to be a risk factor for cognitive decline in Alzheimer’s disease. On the other hand, betaine has been demonstrated to attenuate Alzheimer-like pathological changes induced by homocysteine. It is reasonable to conclude that this is due to triggering the remethylation pathway mediated by betaine-homocysteine-methyltransferase. In the present study, we used the transgenic Caenorhabditis elegans strain CL2006, to test whether betaine is able to reduce β-amyloid-induced paralysis in C. elegans. This model expresses human β-amyloid 1–42 under control of a muscle-specific promoter that leads to progressive, age-dependent paralysis in the nematodes.

Results

Betaine at a concentration of 100 μM was able to reduce homocysteine levels in the presence and absence of 1 mM homocysteine. Simultaneously, betaine both reduced normal paralysis rates in the absence of homocysteine and increased paralysis rates triggered by addition of homocysteine. Knockdown of cystathionine-β-synthase using RNA interference both increased homocysteine levels and paralysis. Additionally, it prevented the reducing effects of betaine on homocysteine levels and paralysis.

Conclusion

Our studies show that betaine is able to reduce homocysteine levels and β-amyloid-induced toxicity in a C. elegans model for Alzheimer’s disease. This effect is independent of the remethylation pathway but requires the transsulfuration pathway mediated by cystathionine-β-synthase.

     

Response to dietary carbohydrates in European seabass (<Emphasis Type="Italic">Dicentrarchus labrax</Emphasis>) muscle tissue as revealed by NMR-based metabolomics

Introduction

Feed optimization is a key step to the environmental and economic sustainability of aquaculture, especially for carnivorous species. Plant-derived ingredients can contribute to reduce costs and nitrogenous effluents while sparing wild fish stocks. However, the metabolic use of carbohydrates from vegetable sources by carnivorous fish is still not completely understood.

Objectives

We aimed to study the effects of diets with carbohydrates of different digestibilities, gelatinized starch (DS) and raw starch (RS), in the muscle metabolome of European seabass (Dicentrarchus labrax).

Methods

We followed an NMR-metabolomics approach, using two sample preparation procedures, the intact muscle (HRMAS) and the aqueous muscle extracts (¹H NMR), to compare the variations in muscle metabolome between the two diets.

Results

In muscle, multivariate analysis revealed similar metabolome shifts for DS and RS diets, when compared with the control diet. HRMAS of intact muscle, which included both hydrophobic and hydrophilic metabolites, showed increased lipid in DS-fed fish by univariate analysis. Regardless of the nature of the starch, increased glycine and phenylalanine, and decreased proline were observed when compared to the Ctr diet. Combined univariate analysis of intact muscle and aqueous extracts indicated specific diet related changes in lipid and amino acid metabolism, consistent with increased dietary carbohydrate supplementation.

Conclusions

Due to differential sample processing, outputs differ in detail but provide complementary information. After tracing nutritional alterations by profiling fillet components, DS seems to be the most promising alternative to fishmeal-based diets in aquaculture. This approach should be reproducible for other farmed fish species and provide valuable information on nutritional and organoleptic properties of the final product.

     

Quantifying and filtering knowledge generated by literature based discovery

Background

Literature based discovery (LBD) automatically infers missed connections between concepts in literature. It is often assumed that LBD generates more information than can be reasonably examined.

Methods

We present a detailed analysis of the quantity of hidden knowledge produced by an LBD system and the effect of various filtering approaches upon this. The investigation of filtering combined with single or multi-step linking term chains is carried out on all articles in PubMed.

Results

The evaluation is carried out using both replication of existing discoveries, which provides justification for multi-step linking chain knowledge in specific cases, and using timeslicing, which gives a large scale measure of performance.

Conclusions

While the quantity of hidden knowledge generated by LBD can be vast, we demonstrate that (a) intelligent filtering can greatly reduce the number of hidden knowledge pairs generated, (b) for a specific term, the number of single step connections can be manageable, and (c) in the absence of single step hidden links, considering multiple steps can provide valid links.

     

<Superscript>1</Superscript>H NMR metabolic profiling of gastric cancer patients with lymph node metastasis

Introduction

Gastric cancer (GC) is a malignant tumor worldwide. As primary pathway for metastasis, the lymphatic system is an important prognostic factor for GC patients. Although the metabolic changes of gastric cancer have been investigated in extensive studies, little effort focused on the metabolic profiling of lymph node metastasis (LNM)-positive or negative GC patients.

Objectives

We performed ¹H NMR spectrum of GC tissue samples with and without LNM to identify novel potential metabolic biomarkers in the process of LNM of GC.

Methods

¹H NMR-based untargeted metabolomics approach combined with multivariate statistical analyses were used to study the metabolic profiling of tissue samples from LNM-positive GC patients (n?=?40), LNM-negative GC patients (n?=?40) and normal controls (n?=?40).

Results

There was a clear separation between GC patients and normal controls, and 33 differential metabolites were identified in the study. Moreover, GC patients were also well-classified according to LNM-positive or negative. Totally eight distinguishing metabolites were selected in the metabolic profiling of GC patients with LNM-positive or negative, suggesting the metabolic dysfunction in the process of LNM. According to further validation and analysis, especially BCAAs metabolism (leucine, isoleucine, valine), GSH and betaine may be as potential factors of diagnose and prognosis of GC patients with or without LNM.

Conclusion

To our knowledge, this is the first metabolomics study focusing on LNM of GC. The identified distinguishing metabolites showed a promising application on clinical diagnose and therapy prediction, and understanding the mechanism underlying the carcinogenesis, invasion and metastasis of GC.

     

Metabolomic study of the response to cold shock in a strain of <Emphasis Type="Italic">Pseudomonas syringae</Emphasis> isolated from cloud water

Introduction

Active microorganisms have been recently discovered in clouds, thus demonstrating the capacity of microorganisms to exist in harsh environments, including exposure to UV and oxidants, osmotic and cold shocks, etc. It is important to understand how microorganisms respond to and survive such stresses at the metabolic level.

Objectives

The objective of this work is to assess metabolome modulation in a strain of Pseudomonas syringae isolated from cloud water and facing temperature downshift from 17 to 5 °C by identifying key molecules and pathways of the response/adaptation to cold shock.

Methods

Bacterial extracts from suspensions of cells grown at 17 °C and further incubated in microcosms at 5 and 17 °C to mimic cloud conditions were analysed by combining LC-MS and NMR; the results were evaluated in comparison to similar suspensions kept at constant temperature. The differences in the metabolome profiles were deciphered using multivariate statistics (PLS-DA).

Results

Key cold shock biomarkers were observed, including cryoprotectants (trehalose, glucose, glycerol, carnitine, glutamate), antioxidants (glutathione and carnitine) and their precursors, alkaloids (bellendine and slaframine) and metabolites involved in energy metabolism (ATP, carbohydrates). Furthermore, new short peptides (nine dipeptides and a tetrapeptide) were found that have no known function.

Conclusions

This study shows that in response to cold temperatures, Pseudomonas syringae PDD-32b-74 demonstrates numerous metabolism modifications to counteract the impacts of low temperatures.

     

Elevated CO<Subscript>2</Subscript> and virus infection impacts wheat and aphid metabolism

Introduction

The aphid Rhopalosiphum padi L. is a vector of Barley yellow dwarf virus (BYDV) in wheat and other economically important cereal crops. Increased atmospheric CO₂ has been shown to alter plant growth and metabolism, enhancing BYDV disease in wheat. However, the biochemical influences on aphid metabolism are not known.

Objectives

This work aims to determine whether altered host-plant quality, influenced by virus infection and elevated CO₂, impacts aphid weight and metabolism.

Methods

Untargeted ¹H NMR metabolomics coupled with multivariate statistics were employed to profile the metabolism of R. padi reared on virus-infected and non-infected (sham-inoculated) wheat grown under ambient CO₂ (aCO₂, 400 µmol mol^?1) and future, predicted elevated CO₂ (eCO₂, 650 µmol mol^?1) concentrations. Un-colonised wheat was also profiled to observe changes to host-plant quality (i.e., amino acids and sugars).

Results

The direct impacts of virus or eCO₂ were compared. Virus presence increased aphid weight under aCO₂ but decreased weight under eCO₂; whilst eCO₂ increased non-viruliferous (sham) aphid weight but decreased viruliferous aphid weight. Discriminatory metabolites due to eCO₂ were succinate and sucrose (in sham wheat), glucose, choline and betaine (in infected wheat), and threonine, lactate, alanine, GABA, glutamine, glutamate and asparagine (in aphids), irrespective of virus presence. Discriminatory metabolites due to virus presence were alanine, GABA, succinate and betaine (in wheat) and threonine and lactate (in aphids), irrespective of CO₂ treatment.

Conclusion

This study confirms that virus and eCO₂ alter host-plant quality, and these differences are reflected by aphid weight and metabolism.

     

Cinnamon: does it hold its promises in cows? Using non-targeted blood serum metabolomics profiling to test the effects of feeding cinnamon to dairy cows undergoing lactation-induced insulin resistance

Introduction

Cinnamon exerts insulin-enhancing activity in vitro and was demonstrated to improve blood glucose and lipid profiles in several human studies. Such effects may have an impact on metabolically stressed cows.

Objective

To study the effects of cinnamon supplementation during the transition from late pregnancy to early lactation on the metabolism in dairy cows.

Methods

Twenty-four Holstein cows (n?=?8/group) were assigned to either the control group (CTR; without supplementation) or the supplementation groups [supplemental cinnamon at 20 (LCIN) or 40 (HCIN) g/cow per day (d)] from 28 d before calving until 21 d thereafter. Blood samples were assayed for glucose, nonesterified fatty acids (NEFA), β-hydroxybutyrate (BHBA), and insulin; an index estimating insulin sensitivity (RQUICKI) was calculated. The serum metabolome was characterized in the samples collected from d 14 using a non-targeted approach.

Results

The serum concentrations of glucose and insulin did not differ among groups and followed a similar pattern over time. The serum NEFA concentrations were greater in LCIN (d 2, 7, and 14) and HCIN (d 14) than in CTR. On d 14 and 21, LCIN and HCIN had greater serum BHBA concentrations than CTR cows. The top 10 metabolites identified with significantly higher levels in the supplemented than the CTR cows were related to fatty acid metabolism.

Conclusion

The data suggest lipolytic and ketogenic effects of cinnamon supplementation in dairy cows during the transition from late gestation to early lactation. The fatty acid metabolites found elevated in the supplemented cows point towards impaired mitochondrial fatty acid β-oxidation.

     

Crystal structure of <Emphasis Type="Italic">Escherichia coli</Emphasis> protein ybgI,a toroidal structure with a dinuclear metal site

Background

The protein encoded by the gene ybgI was chosen as a target for a structural genomics project emphasizing the relation of protein structure to function.

Results

The structure of the ybgI protein is a toroid composed of six polypeptide chains forming a trimer of dimers. Each polypeptide chain binds two metal ions on the inside of the toroid.

Conclusion

The toroidal structure is comparable to that of some proteins that are involved in DNA metabolism. The di-nuclear metal site could imply that the specific function of this protein is as a hydrolase-oxidase enzyme.

     

Quantitative analysis of tetrahydrofolate metabolites from <Emphasis Type="Italic">clostridium autoethanogenum</Emphasis>

Introduction

Quantification of tetrahydrofolates (THFs), important metabolites in the Wood–Ljungdahl pathway (WLP) of acetogens, is challenging given their sensitivity to oxygen.

Objective

To develop a simple anaerobic protocol to enable reliable THFs quantification from bioreactors.

Methods

Anaerobic cultures were mixed with anaerobic acetonitrile for extraction. Targeted LC–MS/MS was used for quantification.

Results

Tetrahydrofolates can only be quantified if sampled anaerobically. THF levels showed a strong correlation to acetyl-CoA, the end product of the WLP.

Conclusion

Our method is useful for relative quantification of THFs across different growth conditions. Absolute quantification of THFs requires the use of labelled standards.

     

Sex differences in hepatic one-carbon metabolism

Background

There are large differences between men and women of child-bearing age in the expression level of 5 key enzymes in one-carbon metabolism almost certainly caused by the sex hormones. These male-female differences in one-carbon metabolism are greatly accentuated during pregnancy. Thus, understanding the origin and consequences of sex differences in one-carbon metabolism is important for precision medicine.

Results

We have created a mathematical model of hepatic one-carbon metabolism based on the underlying physiology and biochemistry. We use the model to investigate the consequences of sex differences in gene expression. We give a mechanistic understanding of observed concentration differences in one-carbon metabolism and explain why women have lower S-andenosylmethionine, lower homocysteine, and higher choline and betaine. We give a new explanation of the well known phenomenon that folate supplementation lowers homocysteine and we show how to use the model to investigate the effects of vitamin deficiencies, gene polymorphisms, and nutrient input changes.

Conclusions

Our model of hepatic one-carbon metabolism is a useful platform for investigating the mechanistic reasons that underlie known associations between metabolites. In particular, we explain how gene expression differences lead to metabolic differences between males and females.

     

Extraction of natural products from bark of <Emphasis Type="Italic">Betula pendula</Emphasis> using ionic liquids

Introduction

Aqueous–methanol mixtures have successfully been applied to extract a broad range of metabolites from plant tissue. However, a certain amount of material remains insoluble.

Objectives

To enlarge the metabolic compendium, two ionic liquids were selected to extract the methanol insoluble part of trunk from Betula pendula.

Methods

The extracted compounds were analyzed by LC/MS and GC/MS.

Results

The results show that 1-butyl-3-methylimidazolium acetate (IL-Ac) predominantly resulted in fatty acids, whereas 1-ethyl-3-methylimidazolium tosylate (IL-Tos) mostly yielded phenolic structures. Interestingly, bark yielded more ionic liquid soluble metabolites compared to interior wood.

Conclusion

From this one can conclude that the application of ionic liquids may expand the metabolic snapshot.

     

Iridoid and phenylethanoid/phenylpropanoid metabolite profiles of <Emphasis Type="Italic">Scrophularia</Emphasis> and <Emphasis Type="Italic">Verbascum</Emphasis> species used medicinally in North America

Introduction

Botanicals containing iridoid and phenylethanoid/phenylpropanoid glycosides are used worldwide for the treatment of inflammatory musculoskeletal conditions that are primary causes of human years lived with disability, such as arthritis and lower back pain.

Objectives

We report the analysis of candidate anti-inflammatory metabolites of several endemic Scrophularia species and Verbascum thapsus used medicinally by peoples of North America.

Methods

Leaves, stems, and roots were analyzed by ultra-performance liquid chromatography-mass spectrometry (UPLC-MS) and partial least squares-discriminant analysis (PLS-DA) was performed in MetaboAnalyst 3.0 after processing the datasets in Progenesis QI.

Results

Comparison of the datasets revealed significant and differential accumulation of iridoid and phenylethanoid/phenylpropanoid glycosides in the tissues of the endemic Scrophularia species and Verbascum thapsus.

Conclusions

Our investigation identified several species of pharmacological interest as good sources for harpagoside and other important anti-inflammatory metabolites.

     

Chilling slows anaerobic metabolism to improve anoxia tolerance of insects

Background

Insects are renowned for their ability to survive anoxia. Anoxia tolerance may be enhanced during chilling through metabolic suppression.

Aims

Here, the metabolomic response of insects to anoxia, both with and without chilling, for different durations (12–36 h) was examined to assess the potential cross-tolerance mechanisms.

Results

Chilling during anoxia (cold anoxia) significantly improved survival relative to anoxia at warmer temperatures. Reduced intermediate metabolites and increased lactic acid, indicating a switch to anaerobic metabolism, were characteristic of larvae in anoxia.

Conclusions

Anoxia tolerance was correlated survival improvements after cold anoxia were correlated with a reduction in anaerobic metabolism.

     

<Emphasis Type="Italic">massPix</Emphasis>: an R package for annotation and interpretation of mass spectrometry imaging data for lipidomics

Introduction

Mass spectrometry imaging (MSI) experiments result in complex multi-dimensional datasets, which require specialist data analysis tools.

Objectives

We have developed massPix—an R package for analysing and interpreting data from MSI of lipids in tissue.

Methods

massPix produces single ion images, performs multivariate statistics and provides putative lipid annotations based on accurate mass matching against generated lipid libraries.

Results

Classification of tissue regions with high spectral similarly can be carried out by principal components analysis (PCA) or k-means clustering.

Conclusion

massPix is an open-source tool for the analysis and statistical interpretation of MSI data, and is particularly useful for lipidomics applications.

     

How the chemical features of molecules may have addressed the settlement of metabolic steps

Introduction

While the evolutionary adaptation of enzymes to their own substrates is a well assessed and rationalized field, how molecules have been originally selected in order to initiate and assemble convenient metabolic pathways is a fascinating, but still debated argument.

Objectives

Aim of the present study is to give a rationale for the preferential selection of specific molecules to generate metabolic pathways.

Methods

The comparison of structural features of molecules, through an inductive methodological approach, offer a reading key to cautiously propose a determining factor for their metabolic recruitment.

Results

Starting with some commonplaces occurring in the structural representation of relevant carbohydrates, such as glucose, fructose and ribose, arguments are presented in associating stable structural determinants of these molecules and their peculiar occurrence in metabolic pathways.

Conclusions

Among other possible factors, the reliability of the structural asset of a molecule may be relevant or its selection among structurally and, a priori, functionally similar molecules.

     

Digestive-resistant carbohydrates affect lipid metabolism in rats

Introduction

Digestion resistant carbohydrates (DRC) are complex carbohydrates that resist digestion and absorption in the small bowel. Diets high in DRC can have wide ranging impacts on the health of the host, which include changes to immunity and allergy, incidence of cardiovascular disease, and obesity.

Objectives

The aim of this study was to characterise the effects of DRC (inulin, konjac or resistant starch) on large intestinal short-chain fatty acid (SCFA) concentrations and serum metabolite and lipid profiles.

Methods

A rat model was used to compare the effects of feeding a basal diet or the basal diet containing 5 % inulin, konjac or resistant starch for 14 days.

Results

Of the three DRC, inulin had the greatest effect; ten serum phospholipids differed significantly in abundance between inulin-treated and control rats. In particular phosphatidylcholines and lysophosphatidylcholines containing fatty acyl chains 22:5, 22:4, 20:4, 18:0 and 16:0 were increased in the inulin-fed group, whereas phosphocholines containing fatty acyls 20:5 and 22:6 were decreased.

Conclusion

These results indicated an impact on both n-3 and n-6 fatty acid metabolism as a result of inulin dietary intake. Increased intestinal concentrations of SCFA were detected in rats fed DRC, but only inulin caused appreciable changes to serum lipid profiles.

     

Association of mitochondrial dysfunction and lipid metabolism with type 2 diabetes mellitus: A review of literature

Background

Diabetes mellitus (DM) is one of the most prevalent chronic diseases, and its prevalence continues to increase globally. The impact of mitochondrial dysfunction and lipid metabolism on diabetes mellitus and insulin resistance (IR) has been implicated in several previous reports; however, the results of studies are confusing despite four decades of study.

Methods/Results

This review has evaluated updated understanding of the role of mitochondrial dysfunction and lipid metabolism on type 2 diabetes, and found that mitochondrial dysfunction and lipid metabolism disorder induce the dysregulation of liver and pancreatic beta cells, insulin resistance, and type 2 diabetes.

Conclusion

Mitochondrial dysfunction and lipid metabolism induce metabolic dysregulation and finally increasing the possibility of diabetes.

     

New insights into the mechanistic action of methyldehydrodieugenol B towards <Emphasis Type="Italic">Leishmania</Emphasis> (L.) <Emphasis Type="Italic">infantum</Emphasis> via a multiplatform based untargeted metabolomics approach

Introduction

Leishmaniasis is a parasitic neglected disease affecting millions of people worldwide. Clinical practice resorts to long and costly treatments with a therapeutic arsenal limited to highly toxic drugs, often associated to adverse side effects. Additionally, resistant strains are reported to be increasing.

Aim

In this work, the mechanistic action of a drug candidate (methydehydrodieugenol B), isolated from twigs of Nectandra leucantha, towards Leishmania infantum was studied by a global metabolomics approach using GC-MS and RPLC-MS platforms.

Method

L. infantum promastigotes were grown in culture medium for 72 h and treated with methydehydrodieugenol B at 58.18 μg.mL-1 concentration; after 48 h treatment, enzyme activity was quenched, cells washed and frozen until analysis. For GC-MS analysis (Fiehn’s method), 1:1 methanol:water extracts were prepared and derivatized with O-methoxyamine in pyridine at room temperature for 90 min, followed by silylation with BSTFA/1% TMCS at 40 °C for 30 min. Pure methanolic extracts were also prepared and analyzed directly by RPLC-MS with a acetonitrile/water mobile phase acidulated with formic acid and gradient elution.

Result

Several amino acids, fatty acids, carbohydrates, and glycerolipids were found as discriminant metabolites, mostly decreased in treated samples. Due to the complexity of the parasite metabolism and the great diversity of altered metabolites, a multi-target mechanism was assigned to the drug candidate, where changes in the cell energy sources and in the lipid composition of the parasite plasma membrane were prominent.

Conclusion

These results contributed to elucidate the broad action of methyldehydrodieugenol B against Leishmania, paving the way in the search of novel alternative therapies.