cis-Aconitic acid

Osmotic Stress Leads to Significant Changes in Rice Root Metabolic Profiles between Tolerant and Sensitive Genotypes.[Pubmed:33172058]

Plants (Basel). 2020 Nov 6;9(11). pii: plants9111503.

To breed osmotic stress-tolerant rice, the mechanisms involved in maintaining root growth under osmotic stress is important to elucidate. In this study, two rice (Oryza sativa L.) cultivars, IR 58 (stress-tolerant cultivar) and Basilanon (stress-sensitive cultivar), were used. After 1, 3, and 7 days of -0.42 MPa osmotic stress treatment induced by polyethylene glycol (PEG) 6000, root metabolomes were analyzed, yielding 276 detected compounds. Among 276 metabolites, 102 metabolites increased with the duration of the stress treatment in IR 58 roots, and only nine metabolites decreased. In contrast, 51 metabolites increased, and 45 metabolites decreased in Basilanon roots. Principal component analysis (PCA) scores clearly indicated differences between the cultivars and the treatments. Pathway analysis showed that the metabolites exhibiting stress-induced increases in IR 58 were those involved in sugar metabolism (such as sucrose 6'-phosphate, glucose 1-phosphate), polyamine and phenylpropanoid metabolisms (such as spermine, spermidine, gamma-aminobutyric acid (GABA)), and glutathione metabolism (such as glutathione, cysteine, cadaverine). IR 58 roots showed an increase in the most proteinogenic amino acids such as proline, serine, glutamine and asparagine. It was also maintained or increased the tricarboxylic acid (TCA) cycle intermediates (citric acid, cis-Aconitic acid, isocitric acid, fumaric acid, malic acid) under osmotic stress compared with that under control. Therefore, IR 58 actively synthesized various metabolites, and the increase in these metabolites contributed to the maintenance of important biological functions such as energy production and antioxidant defense to promote root development under osmotic stress.

Urine Metabolomics Analysis in Patients With Normoalbuminuric Diabetic Kidney Disease.[Pubmed:33123032]

Front Physiol. 2020 Oct 6;11:578799.

Objective: Diabetic kidney disease (DKD) leads to low high albuminuria and gradually progresses to very high albuminuria with kidney insufficiency. However, about 20-40% of DKD is normoalbuminuric DKD (NADKD), which has impaired kidney function but normal urine albumin. This study is to investigate the urine metabolomic profiles of patients with NADKD and albuminuria DKD (ADKD). Methods: In total, 95 patients were divided into a simple diabetes mellitus group (SDM group), an ADKD group, and a NADKD group. All subjects were analyzed for urine metabolites using non-targeted metabolomics based on ultra-performance liquid chromatography - tandem mass spectrometry. Results: The urine metabolomic profiles of the SDM group, NADKD group, and ADKD group were significantly different, and 65 different metabolites were identified among the three groups. Metabolic pathway analysis of these differential metabolites found that the top three significantly changed metabolic pathways were linoleic acid metabolism, citrate cycle, and, arginine and proline metabolism. There are 12 metabolites enriched in these three metabolic pathways. In detail, compared with those in the SDM group, the levels of gamma-linolenic acid in the ADKD group were increased significantly, while the levels of succinic acid, cis-Aconitic acid, citric acid, L-proline, L-erythro-4-hydroxyglutamate, N-methylhydantoin, N-carbamoylputrescine, spermidine, and 5-aminopentanoic acid were reduced significantly; compared with those in the NADKD group, the levels of linoleic acid, gamma-linolenic acid, and L-malic acid in the ADKD group were increased significantly (P < 0.05), while the levels of L-proline, L-erythro-4-hydroxyglutamate, N-carbamoylputrescine, and spermidine were significantly reduced (P < 0.05). However, there were no significant difference between the SDM group and NADKD group (P > 0.05). Conclusion: The urine metabolomic profiles between the NADKD group and the ADKD group are significantly different. Specifically, these two groups have distinct levels of linoleic acid, gamma-linolenic acid, L-malic acid, L-proline, L-erythro-4-hydroxyglutamate, N-carbamoylputrescine, and spermidine.

Exploring the biomarkers associated with different host inflammation of acute respiratory distress syndrome (ARDS) from lung metabolomics in mice.[Pubmed:33049802]

Rapid Commun Mass Spectrom. 2020 Oct 13:e8971.

RATIONALE: The aim of this study was to analyze the metabolomics of lung with different host inflammation of acute respiratory distress syndrome (ARDS) for the identification of biomarkers for predicting severity under different inflammatory conditions. METHODS: Cecal ligation and puncture (CLP) and lipopolysaccharide (LPS)-intratracheal injection induced acute lung injury (ALI). A mouse model was used to explore lung metabolomic biomarkers in ALI/ARDS. The splenectomy model was used as an auxiliary method to distinguish between hyper- and hypo-inflammatory subtypes. Plasma, lung tissue and bronchoalveolar lavage fluid (BALF) samples were collected from mice after CLP/LPS. The severity of lung injury was evaluated. Expression of Tumor Necrosis Factor-alpha (TNF-alpha) in mice serum and lung was tested by ELISA and PCR. Polymorphonuclear cells in BALF were counted. The lung metabolites were detected by GC/MS, and the metabolic pathways predicted using the KEGG database. RESULTS: The LPS/CLP-Splen group had more severe lung injury than the corresponding ALI group; that in the CLP-Splen group was more serious than in the LPS-Splen group. TNF-alpha expression was significantly elevated in the serum and lung tissue after LPS or CLP, and higher in the LPS/CLP-Splen group than in the corresponding ALI group. The level of TNF-alpha in the CLP-Splen group was elevated significantly over that in the LPS-Splen group. Both these groups also showed significant neutrophil exudation within the lungs. During differential inflammation, more differential metabolites were detected in the lungs of the CLP-group ALI mice than inthe LPS group. A total of 41 compounds were detected in the lungs of the CLP and CLP-Splen groups. Contrastingly, 8 compounds were detected in the lungs of the LPS and LPS-Splen groups. The LPS-Splen and CLP-Splen groups had significant neutrophil exudation in the lung. Random forest analysis of lung-targeted metabolomics data indicated 4-hydroxyphenylacetic acid,1-aminocyclopentanecarboxylic acid (ACPC), cis-Aconitic acid, and hydroxybenzoic acid as strong predictors of hyper-inflammatory subgroup in the CLP group. Furthermore, with splenectomy, 13 differential metabolic pathways between the CLP and LPS groups were revealed. CONCLUSIONS: Hyper-inflammatory subgroups of ARDS have a greater inflammatory response and a more active lung metabolism. Combined with host inflammation background, biomarkers from metabolomics could help evaluate the response severity of ARDS.

Constitutive production of aconitate isomerase by Pseudomonas sp. WU-0701 in relation to trans-aconitic acid assimilation.[Pubmed:32994133]

J Biosci Bioeng. 2020 Sep 26. pii: S1389-1723(20)30335-2.

Aconitic acid, an unsaturated tricarboxylic acid, is used in the chemical industry as raw materials for organic synthesis, especially as a specific substrate for a flavoring agent. trans-Aconitic acid (tAA) is a trans-isomer of cis-Aconitic acid and detected in some plants and bacteria. However, biosynthetic route and metabolism of tAA in relation to assimilation have been unknown. Aconitate isomerase (AI; EC 5.3.3.7) catalyzes the reversible isomerization between cis-Aconitic acid and tAA. Pseudomonas sp. WU-0701 was isolated as a bacterium assimilating tAA as sole carbon source, and characterization and gene identification of AI were already reported. Here, we describe that Pseudomonas sp. WU-0701 exhibited growth in each synthetic medium containing glucose, citric acid, isocitric acid, or tAA as sole carbon source. AI was intracellularly detected all the time during the cultivation of the strain WU-0701 cells, irrespective of the carbon sources; AI activity was detected even in the glucose-grown cells. Through the subcellular fractionation experiments, AI was detected in the periplasmic fraction. This is the first report indicating that a bacterium belonging to the genus Pseudomonas is constitutive for the AI production.

Impaired glucose partitioning in primary myotubes from severely obese women with type 2 diabetes.[Pubmed:32966127]

Am J Physiol Cell Physiol. 2020 Dec 1;319(6):C1011-C1019.

The purpose of this study was to determine whether intramyocellular glucose partitioning was altered in primary human myotubes derived from severely obese women with type 2 diabetes. Human skeletal muscle cells were obtained from lean nondiabetic and severely obese Caucasian females with type 2 diabetes [body mass index (BMI): 23.6 +/- 2.6 vs. 48.8 +/- 1.9 kg/m(2), fasting glucose: 86.9 +/- 1.6 vs. 135.6 +/- 12.0 mg/dL, n = 9/group]. 1-[(14)C]-Glucose metabolism (glycogen synthesis, glucose oxidation, and nonoxidized glycolysis) and 1- and 2-[(14)C]-pyruvate oxidation were examined in fully differentiated myotubes under basal and insulin-stimulated conditions. Tricarboxylic acid cycle intermediates were determined via targeted metabolomics. Myotubes derived from severely obese individuals with type 2 diabetes exhibited impaired insulin-mediated glucose partitioning with reduced rates of glycogen synthesis and glucose oxidation and increased rates of nonoxidized glycolytic products, when compared with myotubes derived from the nondiabetic individuals (P < 0.05). Both 1- and 2-[(14)C]-pyruvate oxidation rates were significantly blunted in myotubes from severely obese women with type 2 diabetes compared with myotubes from the nondiabetic controls. Lastly, concentrations of tricarboxylic acid cycle intermediates, namely, citrate (P < 0.05), cis-Aconitic acid (P = 0.07), and alpha-ketoglutarate (P < 0.05), were lower in myotubes from severely obese women with type 2 diabetes. These data suggest that intramyocellular insulin-mediated glucose partitioning is intrinsically altered in the skeletal muscle of severely obese women with type 2 diabetes in a manner that favors the production of glycolytic end products. Defects in pyruvate dehydrogenase and tricarboxylic acid cycle may be responsible for this metabolic derangement associated with type 2 diabetes.

Mitochondrial metabolic substrate utilization in granulosa cells reflects body mass index and total follicle stimulating hormone dosage in in vitro fertilization patients.[Pubmed:32935173]

J Assist Reprod Genet. 2020 Nov;37(11):2743-2756.

PURPOSE: To utilize a novel mitochondrial function assay with pooled granulosa cells to determine whether mitochondrial function would differ by patient demographics and embryo development. METHODS: This was a prospective pilot study in a hospital-based assisted reproductive program and public university. Mitochondrial metabolic substrate utilization was assessed in pooled granulosa cells from 40 women undergoing in vitro fertilization during 2018 and 2019. RESULTS: Assessment of mitochondrial substrate metabolism in pooled granulosa cells revealed higher citric acid, L-malic acid, and octanoyl-L-carnitine utilization with higher body mass index (BMI). Utilization of citric acid, cis-Aconitic acid, D-alpha-keto-glutaric acid, L-glutamine, and alanine plus glycine was significantly lower as total dosage of FSH administered increased. Utilization of glycogen was significantly higher in patients with a higher percentage of fertilized oocytes. D-alpha-keto-glutaric acid utilization was significantly lower in patients with a higher percentage of good 8-cell embryos. L-glutamine utilization was significantly lower, with a higher percentage of blastocyst formation. Mitochondrial metabolic scores (MMS), which reflect overall mitochondrial activity of the granulosa pool, were significantly higher in patients with higher BMI and with greater numbers of mature oocytes retrieved. MMS in granulosa decreased as total FSH dose administered increased. CONCLUSIONS: Granulosa cell utilization of substrates feeding into the citric acid cycle changed with total FSH dosage and BMI. Fertilization rate, 8-cell embryo quality, and blastocyst formation also associated with different energy substrate usage. Mitochondrial substrate utilization by granulosa cells from individual follicles could be further developed into a useful diagnostic tool.

Effect of coexisting manganese ion on the formation of haloacetic acids during chlorination.[Pubmed:32814132]

Chemosphere. 2021 Jan;263:127862.

Haloacetic acids (HAAs) are a group of disinfection by-products formed by the reaction of dissolved organic matter (DOM) in source water and disinfectants in the drinking water treatment process. The formation of HAAs is known to be affected by several factors (e.g., pH, temperature, concentration, and DOM components in source water). However, the effects of coexisting substances, such as metal ions, on HAA formation are not well understood. In this study, HAA formation potentials (FPs) of model compounds of DOM and environmental waters in the presence or absence of manganese ion upon chlorination were compared. The results of experiments with model compounds of DOM showed that manganese ion promoted the formation of HAA from citric acid, trans-aconitic acid, and cis-Aconitic acid. Even for a manganese concentration of less than 50 mug/L, which is the standard value of manganese in drinking water in the USA, EU, and Japan, manganese had great influence on the dichloroacetic acid FPs of these compounds. However, the manganese ion did not enhance the HAAFPs of the environmental waters tested. Nevertheless, manganese may have an effect on HAAFPs of environmental waters collected at the occurrence of an unusual growth of microorganisms, such as algal bloom.

Immune-responsive gene 1 (IRG1) and dimethyl itaconate are involved in the mussel immune response.[Pubmed:32798695]

Fish Shellfish Immunol. 2020 Nov;106:645-655.

Immune-responsive gene 1 (irg1) is a gene that is well-conserved among different taxa and is highly expressed in the mussel Mytilus galloprovincialis at the constitutive level. The expression of this gene increases after a bacterial infection, primarily in haemocytes. irg1 catalyses the production of itaconic acid from cis-Aconitic acid in the Krebs cycle. Recently, itaconate has been revealed as an immune metabolite involved in macrophage polarization. In this work, we studied the effects of exogenous dimethyl itaconate (DI) on mussels in vitro and in vivo at relevant previously described endogenous concentrations and in mussels infected with Vibrio splendidus. DI did not have adverse effects on the haemocytes viability, apoptotic cells, proliferation and phagocytic activity; however, haemocyte size, velocity and accumulated distance were decreased. The antibacterial activity of DI in vitro and in vivo was observed with high concentrations of DI, that is, 30 and 50 mM, respectively. Furthermore, DI inhibited total ROS, increased mitochondrial ROS and modulated antioxidant genes, such as SOD and CAT, related to Nrf2 activation. In this research, we have demonstrated some important pathways in haemocytes in which itaconate can be involved after its production in a bacterial infection.

Assessment of ethanol tolerance of Kluyveromyces marxianus CCT 7735 selected by adaptive laboratory evolution.[Pubmed:32676708]

Appl Microbiol Biotechnol. 2020 Sep;104(17):7483-7494.

Kluyveromyces marxianus CCT 7735 shows potential for producing ethanol from lactose; however, its low ethanol tolerance is a drawback for its industrial application. The first aim of this study was to obtain four ethanol-tolerant K. marxianus CCT 7735 strains (ETS1, ETS2, ETS3, and ETS4) by adaptive laboratory evolution. The second aim was to select among them the strain that stood out and to evaluate metabolic changes associated with the improved ethanol tolerance in this strain. The ETS4 was selected for displaying a specific growth rate higher than the parental strain under ethanol stress (122%) and specific ethanol production rate (0.26 g/g/h) higher than those presented by the ETS1 (0.22 g/g/h), ETS2 (0.17 g/g/h), and ETS3 (0.17 g/g/h) under non-stress condition. Further analyses were performed with the ETS4 in comparison with its parental strain in order to characterize metabolic changes. Accumulation of valine and metabolites of the citric acid cycle (isocitric acid, citric acid, and cis-Aconitic acid) was observed only in the ETS4 subjected to ethanol stress. Their accumulation in this strain may have been important to increase ethanol tolerance. Furthermore, the contents of fatty acid methyl esters and ergosterol were higher in the ETS4 than in the parental strain. These differences likely contributed to enhance ethanol tolerance in the ETS4. KEY POINTS: * K. marxianus ethanol-tolerant strains were selected by adaptive laboratory evolution. * Valine and metabolites of the TCA cycle were accumulated in the ETS4. * High contents of fatty acids and ergosterol contributed to enhance ethanol tolerance.

Metabolomics study of polysaccharide extracts from Polygonatum sibiricum in mice based on (1) H NMR technology.[Pubmed:32424844]

J Sci Food Agric. 2020 Sep;100(12):4627-4635.

BACKGROUND: Polygonatum sibiricum Liliaceae perennial herb, as a commonly used medicine and food homologous plant, has been widely used in clinical practice of Chinese medicine since ancient times, with a history of 2000 years. As the main active ingredient, P. sibiricum polysaccharides have important pharmacological effects in blood sugar reduction and antitumor, antioxidant and liver protection. RESULTS: Mouse models of P. sibiricum polysaccharides were used in combination with (1) H NMR to investigate the metabolic regulation mechanism in mouse tissue and blood. The metabolite maps of the control group and the drug group in the liver had significant changes. The main differential metabolites were glucose 6-phosphate, inositol, lactose, glutamylglycine, galactose, rhamnose, cis-Aconitic acid and histidine, indicating that there was definite correlation between the metabolic detection based on (1) H NMR and the metabolic characteristics of P. sibiricum. The common differential metabolites obtained by overall metabolism analysis were 3-hydroxybutyric acid, d-ribose, adenosine phosphate, inositol, fructose 6-phosphate, histidine, aspartic acid and cis-Aconitic acid. CONCLUSIONS: This work forms the basis for identification of metabolic states combined with metabolic pathways, which could be used as diagnostic and prognostic indicators, providing therapeutic targets for new diseases. (c) 2020 Society of Chemical Industry.

Metabolic engineering of an acid-tolerant yeast strain Pichia kudriavzevii for itaconic acid production.[Pubmed:32346511]

Metab Eng Commun. 2020 Feb 11;10:e00124.

Itaconic acid (IA), or 2-methylenesuccinic acid, has a broad spectrum of applications in the biopolymer industry owing to the presence of one vinyl bond and two acid groups in the structure. Its polymerization can follow a similar mechanism as acrylic acid but additional functionality can be incorporated into the extra beta acid group. Currently, the bio-based production of IA in industry relies on the fermentation of the filamentous fungus Aspergillus terreus. However, the difficulties associated with the fermentation undertaken by filamentous fungi together with the pathogenic potential of A. terreus pose a serious challenge for industrial-scale production. In recent years, there has been increasing interest in developing alternative production hosts for fermentation processes that are more homogenous in the production of organic acids. Pichia kudriavzevii is a non-conventional yeast with high acid tolerance to organic acids at low pH, which is a highly desirable trait by easing downstream processing. We introduced cis-Aconitic acid decarboxylase gene (cad) from A. terreus (designated At_cad) into this yeast and established the initial titer of IA at 135 +/- 5 mg/L. Subsequent overexpression of a native mitochondrial tricarboxylate transporter (herein designated Pk_mttA) presumably delivered cis-aconitate efficiently to the cytosol and doubled the IA production. By introducing the newly invented CRISPR-Cas9 system into P. kudriavzevii, we successfully knocked out both copies of the gene encoding isocitrate dehydrogenase (ICD), aiming to increase the availability of cis-aconitate. The resulting P. kudriavzevii strain, devoid of ICD and overexpressing Pk_mttA and At_cad on its genome produced IA at 505 +/- 17.7 mg/L in shake flasks, and 1232 +/- 64 mg/L in fed-batch fermentation. Because the usage of an acid-tolerant species does not require pH adjustment during fermentation, this work demonstrates the great potential of engineering P. kudriavzevii as an industrial chassis for the production of organic acid.

Metabolomic Profiling of the Aqueous Humor in Patients with Wet Age-Related Macular Degeneration Using UHPLC-MS/MS.[Pubmed:32293180]

J Proteome Res. 2020 Jun 5;19(6):2358-2366.

Assessing metabolomic alterations in age-related macular degeneration (AMD) can provide insights into its pathogenesis. We compared the metabolomic profiles of the aqueous humor between wet AMD patients (n = 26) and age- and sex-matched patients undergoing cataract surgery without AMD as controls (n = 20). A global untargeted metabolomics study was performed using ultra-high-performance liquid chromatography tandem mass spectrometry. Univariate analysis after the false discovery correction showed 18 significantly altered metabolites among the 291 metabolites measured. These differential metabolomic profiles pointed to three interconnected metabolic pathways: a compromised carnitine-associated mitochondrial oxidation pathway (carnitine, deoxycarnitine, N6-trimethyl-l-lysine), an altered carbohydrate metabolism pathway (cis-Aconitic acid, itaconatic acid, and mesaconic acid), which plays a role in senescence and immunity, and an activated osmoprotection pathway (glycine betaine, creatine), which potentially contributes to the pathogenesis of the disease. These results suggested that metabolic dysfunction in AMD is mitochondrial-centered and may provide new insights into the pathophysiology of wet AMD and novel therapeutic strategies.