Microarray and metabolome analysis of hepatic response to fasting and subsequent refeeding in zebrafish (Danio rerio)

Abstract Background Compensatory growth refers to the phenomenon in which organisms grow faster after the improvement of an adverse environment and is thought to be an adaptive evolution to cope with the alleviation of the hostile environment. Many fish have the capacity for compensatory growth, but the underlying cellular mechanisms remain unclear. In the present study, microarray and nontargeted metabolomics were performed to characterize the transcriptome and metabolome of zebrafish liver during compensatory growth. Results Zebrafish could regain the weight they lost during 3 weeks of fasting and reach a final weight similar to that of fish fed ad libitum when refed for 15 days. When refeeding for 3 days, the liver displayed hyperplasia accompanied with decreased triglyceride contents and increased glycogen contents. The microarray results showed that when food was resupplied for 3 days, the liver TCA cycle (Tricarboxylic acid cycle) and oxidative phosphorylation processes were upregulated, while DNA replication and repair, as well as proteasome assembly were also activated. Integration of transcriptome and metabolome data highlighted transcriptionally driven alterations in metabolism during compensatory growth, such as altered glycolysis and lipid metabolism activities. The metabolome data also implied the participation of amino acid metabolism during compensatory growth in zebrafish liver. Conclusion Our study provides a global resource for metabolic adaptations and their transcriptional regulation during refeeding in zebrafish liver. This study represents a first step towards understanding of the impact of metabolism on compensatory growth and will potentially aid in understanding the molecular mechanism associated with compensatory growth.

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Metabolic characterization of aggressive breast cancer cells exhibiting invasive phenotype: Impact of non-cytotoxic doses of 2-DG on diminishing invasiveness

10.21203/rs.3.rs-36631/v3 ◽

2020 ◽

Author(s):

Mayumi Fujita ◽

Kaori Imadome ◽

Veena Somasundaram ◽

Miki Kawanishi ◽

Kumiko Karasawa ◽

...

Keyword(s):

Breast Cancer ◽

Membrane Potential ◽

Cell Lines ◽

Cancer Metabolism ◽

Tricarboxylic Acid Cycle ◽

Prognostic Significance ◽

Tca Cycle ◽

Metabolic Reprogramming ◽

Cancer Therapies ◽

The Impact

Abstract BackgroundMetabolic reprogramming is being recognized as a fundamental hallmark of cancer, and efforts to identify drugs that can target cancer metabolism are underway. In this study, we used human breast cancer (BC) cell lines and established their invading phenotype (INV) collected from transwell inserts to compare metabolome differences and evaluate prognostic significance of the metabolome in aggressive BC invasiveness. MethodsThe invasiveness of seven human BC cell lines were compared using the transwell invasion assay. Among these, INV was collected from SUM149, which exhibited the highest invasiveness. Levels of metabolites in INV were compared with those of whole cultured SUM149 cells (WCC) using CE-TOFMS. The impact of glycolysis in INV was determined by glucose uptake assay using fluorescent derivative of glucose (2-NBDG), and significance of glycolysis, or tricarboxylic acid cycle (TCA) and electron transport chain (ETC) in the invasive process were further determined in aggressive BC cell lines, SUM149, MDA-MB-231, HCC1937, using invasion assays in the presence or absence of inhibitors of glycolysis, TCA cycle or ETC. ResultsSUM149 INV sub-population exhibited a persistent hyperinvasive phenotype. INV were hyper-glycolytic with increased glucose (2-NBDG) uptake; diminished glucose-6-phosphate (G6P) levels but elevated pyruvate and lactate, along with higher expression of phosphorylated-pyruvate dehydrogenase (pPDH) compared to WCC. Notably, inhibiting of glycolysis with lower doses of 2-DG (1 mM), non-cytotoxic to MDA-MB-231 and HCC1937, was effective in diminishing invasiveness of aggressive BC cell lines. In contrast, 3-Nitropropionic acid (3-NA), an inhibitor of succinate dehydrogenase, the enzyme that oxidizes succinate to fumarate in TCA cycle, and functions as complex II of ETC, had no significant effect on their invasiveness, although levels of TCA metabolites or detection of mitochondrial membrane potential with JC-1 staining, indicated that INV cells originally had functional TCA cycles and membrane potential. ConclusionsHyper-glycolytic phenotype of invading cells caters to rapid energy production required for invasion while TCA cycle/ETC cater to cellular energy needs for sustenance in aggressive BC. Lower, non-cytotoxic doses of 2-DG can hamper invasion and can potentially be used as an adjuvant with other anti-cancer therapies without the usual side-effects associated with cytotoxic doses.

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Metabolome Analysis Revealed the Mechanism of Exogenous Glutathione to Alleviate Cadmium Stress in Maize (Zea mays L.) Seedlings

Plants ◽

10.3390/plants10010105 ◽

2021 ◽

Vol 10 (1) ◽

pp. 105

Author(s):

Runfeng Wang ◽

Kaina Lin ◽

Huabin Chen ◽

Zhenyu Qi ◽

Bohan Liu ◽

...

Keyword(s):

Amino Acid ◽

Shikimic Acid ◽

Tricarboxylic Acid Cycle ◽

Tca Cycle ◽

Antioxidant Systems ◽

Cd Stress ◽

Metabolome Analysis ◽

Induced Stress ◽

Leaves And Roots ◽

Amino Acid Contents

Cadmium (Cd) is one of the major heavy metal pollutants in the environment and imposes severe limitations on crop growth and production. Glutathione (GSH) plays an important role in plant Cd tolerance which is able to scavenge stresses-induced reactive oxygen species (ROS) and is involved in the biosynthesis of phytochelatins (PCs). Our previous study revealed that Cd stress affects maize growth, and the GSH treatment could relieve Cd stress in maize seedlings. In this study, we attempted to characterize the metabolomics changes in maize leaves and roots under Cd stress and exogenous GSH conditions. We identified 145 and 133 metabolites in the leaves and roots, respectively. Cd stress decreased the tricarboxylic acid cycle (TCA cycle) metabolism and increased the amino acid contents in the leaves, while it decreased the amino acid contents, increased the TCA cycle metabolism, the sugar contents, and shikimic acid metabolism in the roots. On the other hand, exogenous GSH increased the GSH content, changed the production of metabolites related to antioxidant systems (such as ascorbic acid-related metabolites and flavonoid-related metabolites), and alleviated lipid peroxidation, thereby alleviating the toxic effect of Cd stress on maize. These findings support the idea that GSH alleviates Cd-induced stress in maize and may help to elucidate the mechanism governing Cd-induced stress and the GSH-driven alleviation effect.

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Metabolic characterization of aggressive breast cancer cells exhibiting invasive phenotype: impact of non-cytotoxic doses of 2-DG on diminishing invasiveness

BMC Cancer ◽

10.1186/s12885-020-07414-y ◽

2020 ◽

Vol 20 (1) ◽

Author(s):

Mayumi Fujita ◽

Kaori Imadome ◽

Veena Somasundaram ◽

Miki Kawanishi ◽

Kumiko Karasawa ◽

...

Keyword(s):

Breast Cancer ◽

Membrane Potential ◽

Cell Lines ◽

Cancer Metabolism ◽

Tricarboxylic Acid Cycle ◽

Prognostic Significance ◽

Tca Cycle ◽

Metabolic Reprogramming ◽

Cancer Therapies ◽

The Impact

Abstract Background Metabolic reprogramming is being recognized as a fundamental hallmark of cancer, and efforts to identify drugs that can target cancer metabolism are underway. In this study, we used human breast cancer (BC) cell lines and established their invading phenotype (INV) collected from transwell inserts to compare metabolome differences and evaluate prognostic significance of the metabolome in aggressive BC invasiveness. Methods The invasiveness of seven human BC cell lines were compared using the transwell invasion assay. Among these, INV was collected from SUM149, which exhibited the highest invasiveness. Levels of metabolites in INV were compared with those of whole cultured SUM149 cells (WCC) using CE-TOFMS. The impact of glycolysis in INV was determined by glucose uptake assay using fluorescent derivative of glucose (2-NBDG), and significance of glycolysis, or tricarboxylic acid cycle (TCA) and electron transport chain (ETC) in the invasive process were further determined in aggressive BC cell lines, SUM149, MDA-MB-231, HCC1937, using invasion assays in the presence or absence of inhibitors of glycolysis, TCA cycle or ETC. Results SUM149 INV sub-population exhibited a persistent hyperinvasive phenotype. INV were hyper-glycolytic with increased glucose (2-NBDG) uptake; diminished glucose-6-phosphate (G6P) levels but elevated pyruvate and lactate, along with higher expression of phosphorylated-pyruvate dehydrogenase (pPDH) compared to WCC. Notably, inhibiting of glycolysis with lower doses of 2-DG (1 mM), non-cytotoxic to MDA-MB-231 and HCC1937, was effective in diminishing invasiveness of aggressive BC cell lines. In contrast, 3-Nitropropionic acid (3-NA), an inhibitor of succinate dehydrogenase, the enzyme that oxidizes succinate to fumarate in TCA cycle, and functions as complex II of ETC, had no significant effect on their invasiveness, although levels of TCA metabolites or detection of mitochondrial membrane potential with JC-1 staining, indicated that INV cells originally had functional TCA cycles and membrane potential. Conclusions Hyper-glycolytic phenotype of invading cells caters to rapid energy production required for invasion while TCA cycle/ETC cater to cellular energy needs for sustenance in aggressive BC. Lower, non-cytotoxic doses of 2-DG can hamper invasion and can potentially be used as an adjuvant with other anti-cancer therapies without the usual side-effects associated with cytotoxic doses.

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CNTF enhances aerobic glycolysis and anabolic activities in degenerating retinas

10.1101/2021.12.21.473752 ◽

2021 ◽

Author(s):

Kun Do Rhee ◽

Yanjie Wang ◽

Johanna ten Hoeve ◽

Linsey Stiles ◽

Thao T.T. Nguyen ◽

...

Keyword(s):

Aerobic Glycolysis ◽

Redox Homeostasis ◽

Chain Complex ◽

Tca Cycle ◽

Neuroprotective Activity ◽

Mitochondrial Morphology ◽

Cellular Mechanisms ◽

Expression Of Genes ◽

The Impact ◽

Complex Activities

Ciliary neurotrophic factor (CNTF) has potent neuroprotective activity in retinal degeneration animal models, yet the cellular mechanisms underlying its broad neuronal survival effects remain unclear. Here, we investigated the impact of CNTF on retinal metabolism in a mouse model of human retinitis pigmentosa. CNTF treatment resulted in improved mitochondrial morphology in mutant rod photoreceptors, but also led to reduced oxygen consumption and suppression of respiratory chain complex activities. Metabolomics analyses detected significantly higher levels of ATP and the energy currency phospho-creatine post CNTF exposure. In addition, CNTF-treated retinas contained elevated glycolytic metabolites and showed increased expression of genes and active enzymes of the glycolytic pathway. Metabolomics analyses also revealed increased TCA cycle products, lipid biosynthetic pathway intermediates, nucleotides, and amino acids, indicating an overall CNTF-dependent augmentation of anabolic activities. Moreover, CNTF treatment restored the key antioxidant glutathione to the wild type level in the degenerating retina. Taken together, these results demonstrate that CNTF profoundly impacts the metabolic status of degenerating retinas by promoting aerobics glycolysis and anabolism, enhancing energy supply, and restoring redox homeostasis. Our study thus reveals important cellular mechanisms underlying CNTF-mediated neuroprotection and provides novel insight for the on-going CNTF clinical trials treating blinding diseases.

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Effect of Huangqin Tang on Urine Metabolic Profile in Rats with Ulcerative Colitis Based on UPLC-Q-Exactive Orbitrap MS

Evidence-based Complementary and Alternative Medicine ◽

10.1155/2020/1874065 ◽

2020 ◽

Vol 2020 ◽

pp. 1-11

Author(s):

Dunfang Wang ◽

Xuran Ma ◽

Shanshan Guo ◽

Yanli Wang ◽

Tao Li ◽

...

Keyword(s):

Ulcerative Colitis ◽

Fatty Acids ◽

Amino Acid ◽

Amino Acid Metabolism ◽

Acid Metabolism ◽

Unsaturated Fatty Acids ◽

Tricarboxylic Acid Cycle ◽

Tca Cycle ◽

Q Exactive ◽

Therapeutic Mechanisms

As a classic prescription, Huangqin Tang (HQT) has been widely applied to treat ulcerative colitis (UC), although its pharmacological mechanisms are not clear. In this study, urine metabolomics was first analysed to explore the therapeutic mechanisms of HQT in UC rats induced by TNBS. We identified 28 potential biomarkers affected by HQT that might cause changes in urine metabolism in UC rats, mapped the network of metabolic pathways, and revealed how HQT affects metabolism of UC rats. The results showed that UC affects amino acid metabolism and biosynthesis of unsaturated fatty acids and impairs the tricarboxylic acid cycle (TCA cycle). UC induced inflammatory and gastrointestinal reactions by inhibiting the transport of fatty acids and disrupting amino acid metabolism. HQT plays key roles via regulating the level of biomarkers in the metabolism of amino acids, lipids, and so on, normalizing metabolic disorders. In addition, histopathology and other bioinformatics analysis further confirm that HQT altered UC rat physiology and pathology, ultimately affecting metabolic function of UC rats.

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An Untargeted Metabolomics Investigation of Milk from Dairy Cows with Clinical Mastitis by 1H-NMR

Foods ◽

10.3390/foods10081707 ◽

2021 ◽

Vol 10 (8) ◽

pp. 1707

Author(s):

Chenglin Zhu ◽

Kaiwei Tang ◽

Xuan Lu ◽

Junni Tang ◽

Luca Laghi

Keyword(s):

Dairy Cows ◽

1H Nmr ◽

Tricarboxylic Acid Cycle ◽

Tca Cycle ◽

Quantitative Information ◽

Clinical Mastitis ◽

Economic Losses ◽

Resonance Spectroscopy ◽

Mastitic Milk ◽

Chemical Groups

Mastitis is one of the diseases with the highest incidence in dairy cows, causing huge economic losses to the dairy industry all over the world. The aim of the study was to characterize mastitic milk metabolome through untargeted nuclear magnetic resonance spectroscopy (1H-NMR). Taking advantage of the high reproducibility of 1H-NMR, we had the opportunity to provide quantitative information for all the metabolites identified. Fifty-four molecules were characterized, sorted mainly into the chemical groups, namely amino acids, peptides and analogues, carbohydrates and derivates, organic acids and derivates, nucleosides, nucleotides and analogues. Combined with serum metabolomic investigations, several pathways were addressed to explain the mechanisms of milk metabolome variation affected by clinical mastitis, such as tricarboxylic acid cycle (TCA cycle) and phenylalanine, tyrosine and tryptophan biosynthesis. These results provide a further understanding of milk metabolome altered by clinical mastitis, which can be used as a reference for the further milk metabolome investigations.

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Production of succinate by engineered strains of Synechocystis PCC 6803 overexpressing phosphoenolpyruvate carboxylase and a glyoxylate shunt

Microbial Cell Factories ◽

10.1186/s12934-021-01529-y ◽

2021 ◽

Vol 20 (1) ◽

Author(s):

Claudia Durall ◽

Kateryna Kukil ◽

Jeffrey A. Hawkes ◽

Alessia Albergati ◽

Peter Lindblad ◽

...

Keyword(s):

Phosphoenolpyruvate Carboxylase ◽

Isocitrate Lyase ◽

Tricarboxylic Acid Cycle ◽

Cell Metabolism ◽

Tca Cycle ◽

Malate Synthase ◽

Glyoxylate Shunt ◽

Control Strain ◽

Genes Encoding ◽

Pcc 6803

Abstract Background Cyanobacteria are promising hosts for the production of various industrially important compounds such as succinate. This study focuses on introduction of the glyoxylate shunt, which is naturally present in only a few cyanobacteria, into Synechocystis PCC 6803. In order to test its impact on cell metabolism, engineered strains were evaluated for succinate accumulation under conditions of light, darkness and anoxic darkness. Each condition was complemented by treatments with 2-thenoyltrifluoroacetone, an inhibitor of succinate dehydrogenase enzyme, and acetate, both in nitrogen replete and deplete medium. Results We were able to introduce genes encoding the glyoxylate shunt, aceA and aceB, encoding isocitrate lyase and malate synthase respectively, into a strain of Synechocystis PCC 6803 engineered to overexpress phosphoenolpyruvate carboxylase. Our results show that complete expression of the glyoxylate shunt results in higher extracellular succinate accumulation compared to the wild type control strain after incubation of cells in darkness and anoxic darkness in the presence of nitrate. Addition of the inhibitor 2-thenoyltrifluoroacetone increased succinate titers in all the conditions tested when nitrate was available. Addition of acetate in the presence of the inhibitor further increased the succinate accumulation, resulting in high levels when phosphoenolpyruvate carboxylase was overexpressed, compared to control strain. However, the highest succinate titer was obtained after dark incubation of an engineered strain with a partial glyoxylate shunt overexpressing isocitrate lyase in addition to phosphoenolpyruvate carboxylase, with only 2-thenoyltrifluoroacetone supplementation to the medium. Conclusions Heterologous expression of the glyoxylate shunt with its central link to the tricarboxylic acid cycle (TCA) for acetate assimilation provides insight on the coordination of the carbon metabolism in the cell. Phosphoenolpyruvate carboxylase plays an important role in directing carbon flux towards the TCA cycle.

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TCA Cycle and Fatty Acids Oxidation Reflect Early Cardiorenal Damage in Normoalbuminuric Subjects with Controlled Hypertension

Antioxidants ◽

10.3390/antiox10071100 ◽

2021 ◽

Vol 10 (7) ◽

pp. 1100

Author(s):

Aranzazu Santiago-Hernandez ◽

Marta Martin-Lorenzo ◽

Ariadna Martin-Blazquez ◽

Gema Ruiz-Hurtado ◽

Maria G Barderas ◽

...

Keyword(s):

Fatty Acids ◽

Tricarboxylic Acid Cycle ◽

Renin Angiotensin System ◽

Roc Curves ◽

Tca Cycle ◽

Urinary Metabolites ◽

Organ Damage ◽

Fatty Acid Binding ◽

Angiotensin System

Moderately increased albuminuria, defined by an albumin to creatinine ratio (ACR) > 30 mg/g, is an indicator of subclinical organ damage associated with a higher risk of cardiovascular and renal disease. Normoalbuminuric subjects are considered at no cardiorenal risk in clinical practice, and molecular changes underlying early development are unclear. To decipher subjacent mechanisms, we stratified the normoalbuminuria condition. A total of 37 hypertensive patients under chronic renin–angiotensin system (RAS) suppression with ACR values in the normoalbuminuria range were included and classified as control (C) (ACR < 10 mg/g) and high-normal (HN) (ACR = 10–30 mg/g). Target metabolomic analysis was carried out by liquid chromatography and mass spectrometry to investigate the role of the cardiorenal risk urinary metabolites previously identified. Besides this, urinary free fatty acids (FFAs), fatty acid binding protein 1 (FABP1) and nephrin were analyzed by colorimetric and ELISA assays. A Mann–Whitney test was applied, ROC curves were calculated and Spearman correlation analysis was carried out. Nine metabolites showed significantly altered abundance in HN versus C, and urinary FFAs and FABP1 increased in HN group, pointing to dysregulation in the tricarboxylic acid cycle (TCA) cycle and fatty acids β-oxidation. We showed here how cardiorenal metabolites associate with albuminuria, already in the normoalbuminuric range, evidencing early renal damage at a tubular level and suggesting increased β-oxidation to potentially counteract fatty acids overload in the HN range.

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Abstract 313: Nonionic Acid is a Target Metabolite of Diabetic Microvasculopathy

Circulation Research ◽

10.1161/res.119.suppl_1.313 ◽

2016 ◽

Vol 119 (suppl_1) ◽

Author(s):

Yasuko K Bando ◽

Haruya Kawase ◽

Kazuyuki Nishimura ◽

Akio Monji ◽

Toyoaki Murohara

Keyword(s):

Diabetic Microangiopathy ◽

Tube Length ◽

Nonanoic Acid ◽

Branch Points ◽

Metabolome Analysis ◽

Whole Heart ◽

The Impact ◽

Type 2 Diabetic

Aim: The purpose of this study is to screen the target metabolites of diabetic microangiopathy in heart by use of whole heart metabolome analysis. Methods: Diet-induced type 2 diabetic mouse were divided into two groups; control and those treated with diabetic remedy GLP-1 receptor agonist Ex-4 for 5 weeks. Mice were euthanized and analyzed at the age of 16 week-old. Results: The capillary density of the T2DM was reduced as compared to those non-diabetic counterparts, which was restored by Ex4 treatment. Changes in angiogenic signals detected by immunoblotting analysis revealed that the phosphorylation levels of eNOS and AMPK were elevated by Ex-4, but those Akt remained unchanged. Tube formation assay revealed that Ex-4 increased tube length and branch points in HUVECs. Consistently with the trend that was observed in vivo experiment, AMPK and eNOS phosphorylation levels were enhanced by Ex4 without affecting Akt phosphorylation. To screen the candidate metabolites that is responsible for the diabetic microvasculopathy in GLP-1-dependent fashion, we performed metabolome analysis by using the whole heart of each mouse. The hierarchical cluster analysis revealed that nonanoic acid (NNA) was the only metabolite that increased in type 2 diabetic mice with concomitant decline by Ex-4 treatment. We next examined the impact of nonanoic acid on in vitro angiogenesis and found that NNA suppressed tube length and branch points in HUVECs in a dose-dependent fashion. Interestingly, NNA canceled eNOS and AMPK phosphorylation that was enhanced by Ex4. Conclusion: GLP-1 ameliorated diabetic microvasculopathy via the AMPK and eNOS axis. NNA is presumably one of the novel anti-angiogenic metabolites that causes diabetic microangiopathy.

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HISTOCHEMICAL INVESTIGATIONS ON THE IN VIVO EFFECTS OF FLUORIDE ON TRICARBOXYLIC ACID CYCLE DEHYDROGENASES FROM PELARGONIUM ZONALE: PART II

Journal of Histochemistry & Cytochemistry ◽

10.1177/15.4.202 ◽

1967 ◽

Vol 15 (4) ◽

pp. 202-206

Author(s):

C. JAMES LOVELACE ◽

GENE W. MILLER

Keyword(s):

Sodium Fluoride ◽

Reductase Activity ◽

Tricarboxylic Acid Cycle ◽

Leaf Tissue ◽

Tca Cycle ◽

Tricarboxylic Acid ◽

Pelargonium Zonale ◽

Acid Cycle ◽

In Vivo Effects

In vivo effects of fluoride on tricarboxylic acid (TCA) cycle dehydrogenase enzymes of Pelargonium zonale were studied using p-nitro blue tetrazoleum chloride. Plants were exposed to 17 ppb HF, and enzyme activities in treated plants were compared to those in controls. Leaves of control plants were incubated in 5 x 10–3 M sodium fluoride. Injuries observed in fumigation and solution experiments were similar. Leaf tissue subjected to HF or sodium fluoride evidenced less succinic p-nitro blue tetrazoleum reductase activity than did control tissue. Other TCA cycle dehydrogenase enzymes were not observably affected by the fluoride concentrations used in these experiments. Excised leaves cultured in 5 x 10–3 M sodium fluoride exhibited less succinic p-nitro blue tetrazoleum reductase activity after 24 hr than did leaves cultured in 5 x 10–3 M sodium chloride.

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