Tricarboxilic Acid Cycle Research Articles

Cholesin receptor signalling is active in cardiovascular system-associated adipose tissue and correlates with SGLT2i treatment in patients with diabetes

BackgroundRecently deorphanized G protein-coupled receptor 146 (GPR146) was shown to respond to signal from a newly identified hormone—cholesin—and to play a role in hepatic lipid metabolism. However, the importance of its biological activity in human organism remains elusive, mainly due to the lack of studies on human tissues up to this point. This study aimed to identify the cholesin receptor-associated genes and clinical factors linked with their expression in cardiovascular system and associated adipose tissues.MethodsRight cardiac auricle, aortic wall, saphenous vein, and adipose tissue (periaortic-PAT, epicardial-EAT, thymic-TAT) samples were collected during coronary artery bypass grafting. Clinical records of the study participants were assessed for the presence of diabetes, medications taken and serum cholesterol levels. GPR146 mRNA expression in all gathered tissues was assessed with qPCR, and RNA seqencing was performed in selected tissues of 20 individuals to identify pathways associated with GPR146 expression.ResultsWe included 46 participants [37 male, 23 with type 2 diabetes, median age 68.50 (Q1–Q3: 63.00–72.00) years, BMI 28.39 (26.06–31.49) kg/m2]. GPR146 expression in adipose tissues significantly correlated with BMI, c-peptide, total cholesterol, and LDL concentrations. Selected metabolic pathways were significantly and positively enriched in GPR146-dependent manner. GPR146-coexpressed genes contained key regulators of lipid metabolism involved in such pathways as fatty acid metabolism, tricarboxilic acid cycle and peroxisomal metabolism. Those genes correlated positively with serum concentrations of LDL, HDL, and total cholesterol. SGLT2i treatment was associated with inversion of GPR146-related signature in EAT, suggesting potential impact on cholesin-GPR146 network.ConclusionsGPR146 expression is associated with serum lipids and metabolically-relevant transcriptomic changes in EAT similar to SGLT2i-associated ones.Graphical abstract

Combined Analysis of the Metabolome and Transcriptome to Explore Heat Stress Responses and Adaptation Mechanisms in Celery (Apium graveolens L.).

Celery is an important leafy vegetable that can grow during the cool season and does not tolerate high temperatures. Heat stress is widely acknowledged as one of the main abiotic stresses affecting the growth and yield of celery. The morphological and physiological indices of celery were investigated in the present study to explore the physiological mechanisms in response to high temperatures. Results showed that the antioxidant enzyme activity, proline, relative conductivity, and malondialdehyde were increased, while chlorophyll and the water content of leaves decreased under high-temperature conditions. Short-term heat treatment increased the stomatal conductance to cool off the leaves by transpiration; however, long-term heat treatment led to stomatal closure to prevent leaf dehydration. In addition, high temperature caused a disordered arrangement of palisade tissue and a loose arrangement of spongy tissue in celery leaves. Combined metabolomic and transcriptomic analyses were further used to reveal the regulatory mechanisms in response to heat stress at the molecular level in celery. A total of 1003 differential metabolites were identified and significantly enriched in amino acid metabolism and the tricarboxilic acid (TCA) cycle. Transcriptome sequencing detected 24,264 different genes, including multiple transcription factor families such as HSF, WRKY, MYB, AP2, bZIP, and bHLH family members that were significantly upregulated in response to heat stress, suggesting that these genes were involved in the response to heat stress. In addition, transcriptional and metabolic pathway analyses showed that heat stress inhibited the glycolysis pathway and delayed the TCA cycle but increased the expression of most amino acid synthesis pathways such as proline, arginine, and serine, consistent with the results of physiological indicators. qRT-PCR further showed that the expression pattern was similar to the expression abundance in the transcriptome. The important metabolites and genes in celery that significantly contributed to the response to high temperatures were identified in the present study, which provided the theoretical basis for breeding heat-resistant celery.

Pre-ischaemic mitochondrial substrate constraint by inhibition of malate-aspartate shuttle preserves mitochondrial function after ischaemia-reperfusion.

Pre-ischaemic administration of aminooxiacetate (AOA), an inhibitor of the malate-aspartate shuttle (MAS), provides cardioprotection against ischaemia-reperfusion injury. The underlying mechanism remains unknown. We examined whether transient inhibition of the MAS during ischaemia and early reperfusion by AOA treatment could prevent mitochondrial damage at later reperfusion. The AOA treatment preserved mitochondrial respiratory capacity with reduced mitochondrial oxidative stress during late reperfusion to the same extent as ischaemic preconditioning (IPC). However, AOA treatment, but not IPC, reduced the myocardial interstitial concentration of tricarboxylic acid cycle intermediates at the onset of reperfusion. The results obtained in the present study demonstrate that metabolic regulation by inhibition of the MAS at the onset of reperfusion may be beneficial for the preservation of mitochondrial function during late reperfusion in an IR-injured heart. Mitochondrial dysfunction plays a central role in ischaemia-reperfusion (IR) injury. Pre-ischaemic administration of aminooxyacetate (AOA), an inhibitor of the malate-aspartate shuttle (MAS), provides cardioprotection against IR injury, although the underlying mechanism remains unknown. We hypothesized that a transient inhibition of the MAS during ischaemia and early reperfusion could preserve mitochondrial function at later phase of reperfusion in the IR-injured heart to the same extent as ischaemic preconditioning (IPC), which is a well-validated cardioprotective strategy against IR injury. In the present study, we show that pre-ischaemic administration of AOA preserved mitochondrial complex I-linked state 3 respiration and fatty acid oxidation during late reperfusion in IR-injured isolated rat hearts. AOA treatment also attenuated the excessive emission of mitochondrial reactive oxygen species during state 3 with complex I-linked substrates during late reperfusion, which was consistent with reduced oxidative damage in the IR-injured heart. As a result, AOA treatment reduced infarct size after reperfusion. These protective effects of MAS inhibition on the mitochondria were similar to those of IPC. Intriguingly, the protection of mitochondrial function by AOA treatment appears to be different from that of IPC because AOA treatment, but not IPC, downregulated myocardial tricarboxilic acid (TCA)-cycle intermediates at the onset of reperfusion. MAS inhibition thus preserved mitochondrial respiratory capacity and decreased mitochondrial oxidative stress during late reperfusion in the IR-injured heart, at least in part, via metabolic regulation of TCA cycle intermediates in the mitochondria at the onset of reperfusion.

A benzimidazole derivative (RCB20) in vitro induces an activation of energetic pathways on Taenia crassiceps (ORF strain) cysticerci

Human cysticercosis caused by Taenia crassiceps is unusual; however, it is an useful experimental model for cysticercosis studies. Benzimidazole derivatives are important antihelminthic drugs widely used against helminths. A novel compound 6-chloro-5-(1-naphthyloxy) -2-(trifluoromethyl)-1H-benzimidazole (RCB20) is a benzimidazole derivative less polar and more lipophilic. The aim of this study was to detect the effect of the RCB20 on the in vitro energetic metabolism of T. crassiceps cysticerci. For this, products of the metabolism both produced and secreted/excreted (S/E) by the parasite were detected through spectrophotometry and high performance liquid chromatography after exposure to 6.5 and 13 μM of RCB20 and albendazole sulfoxide (ABZSO). There was a gradual increase in the concentrations of glucose not uptaken by parasites exposed to both concentrations RCB20 and ABZSO. There was a higher concentration of all the organic acids related to the tricarboxilic acid cycle int the parasites exposed to RCB20. The structural differences between RCB20 and ABZSO result in different targets within the parasite and in a greater induction of the energetic pathways, such as the glycolysis and the TCA cycle. RCB20 is a good candidate as a substitute for anthelminthic benzimidazoles due to a differentiated site of action with similar outcome.

Anoxia in the Domanik basin of the Timan–Pechora region

Analysis of the carbon isotopic composition in aryl carotenoid derivatives, including isorenieratene, in bitumen from domanikoid rocks of the sections cropping out along the Chut River revealed that they contain anomalously heavy carbon, which is determined by the formation of these compounds from green sulfur bacteria of the family Chlorobiaceae. These bacteria use the peculiar process of carbon fixing as HCO3− in the reversed tricarboxilic acid cycle. The fact of hydrosulfuric contamination of the photic water layer in the Domanik basin is confirmed by the plurality of data. Anoxity in shelf water of the middle Frasnian Timan–Pechora basin is evident from lithological features such as, for example, lamination of some rocks and mass death of the fauna.

A microbiological and biogeochemical investigation of the cold seep tubeworm Escarpia southwardae (Annelida: Siboglinidae): Symbiosis and trace element composition of the tube

Tubeworms within the annelid family Siboglinidae rely on sulfur-oxidizing autotrophic bacterial symbionts for their nutrition, and are among the dominant metazoans occurring at deep-sea hydrocarbon seeps. Contrary to their relatives from hydrothermal vents, sulfide uptake for symbionts occurs within the anoxic subsurface sediment, in the posterior ‘root’ region of the animal. This study reports on an integrated microbiological and geochemical investigation of the cold seep tubeworm Escarpia southwardae collected at the Regab pockmark (Gulf of Guinea). Our aim was to further constrain the links between the animal and its symbiotic bacteria, and their environment. We show that E. southwardae harbors abundant sulfur-oxidizing bacterial symbionts in its trophosome. Symbionts are able to fix inorganic carbon using the Calvin-Benson cycle, as reported in most other Siboglinidae, but can also use the reverse Tricarboxilic Acid Cycle. Surprisingly, the observed bacteria appear to be more closely related to symbionts of Escarpia and Lamellibrachia species from very distant sites located in the Gulf of Mexico and eastern Pacific, than to symbionts of a siboglinid occurring at a nearby methane seep site, only a few hundred km away from Regab. Then, by combining scanning electron microscopy and trace element (Mn, Fe, Sr, Zr) analyses of E. southwardae tube, we also show that two distinct oxidation fronts occur along the tube. The first one, near the posterior end of the tube, corresponds to the interface between oxic bottom waters and the underlying anoxic sediment. In contrast, the second redox front is located in the most anterior part of the tube, and could result from active oxygen uptake by the plume of the tubeworm. We speculate that intense oxygen consumption in this region could create favorable conditions for sulfate reduction by specialized bacteria associated with the plume, possibly leading to an additional source of dissolved sulfide that would further enhance the productivity of bacterial symbionts.

Disorders of Primary Metabolites in Response to Drought May Increase the Synthesis of Natural Products for Medicinal Purposes: South American Herbs - a Case Study

The use of plants for healing diseases is one of the oldest medical practices and there are several studies showing that botany and medicine are related. Recent researches have shown that around 25% of new chemical entities and 42% of anticancer drugs marketed worldwide from 1981 to 2006 are obtained from natural products and their derivatives. One-third of the botanical biodiversity of the planet is in South American tropical ecosystems (Neotropical). Over the centuries, plants have been used by indigenous people for curing diseases. This strong tradition has been a great challenge for the scientific community in order to validate the folkloric medicinal use of herbs. In this way, a very few Neotropical pharmaceutical products have reached the market in industrialized countries, even though they have a considerable plant diversity. Plants synthesize several organic compounds which are not related to their growth and development and are called secondary metabolites or natural products that are derived from central or primary metabolism. Because plants are sessile organisms, they have to respond quickly to environmental changes in order to escape and survive under unfavorable conditions. Drought is one of the most worldwide serious impediments for crop yields producing adverse negative effects on plant growth, by impacting leaves and roots growth, stomatal conductance, photosynthetic rate and biomass gain. The three major classes of secondary metabolites are produced from pathways of different primary metabolites, including glycolysis, tricarboxilic acid cycle, aliphatic amino acids, pentose phosphate pathway, shikimate pathway and aromatic amino acids. This review compiles the metabolic changes occurring at primary metabolite level and total biosynthesis of natural products with potential for the development of new drugs in response to drought.

Targeted Metabolomics of Physaria fendleri, an Industrial Crop Producing Hydroxy Fatty Acids

Physaria fendleri (syn. Lesquerella) is a Brassicaceae producing lesquerolic acid, a highly valued hydroxy fatty acid that could be used for several industrial applications, such as cosmetics, lubricating greases, paints, plastics and biofuels. Free of toxins, Physaria oil is an attractive alternative to imported castor (Ricinus communis) oil, and is hence on the verge of commercialization. Gas chromatography-mass spectrometry analysis of fatty acid methyl esters revealed that lesquerolic acid was synthesized and accumulated in the embryos, reaching 60% (w/w) of the total fatty acids. The sequential extraction and characterization of biomass compounds revealed that Physaria embryo metabolism switched from protein to fatty acid biosynthesis between 18 and 24 days post-anthesis (DPA). In order to unravel the metabolic pathways involved in fatty acid synthesis, a targeted metabolomics study was conducted on Physaria embryos at different stages of development. For this purpose, two novel high-throughput liquid chromatography-tandem mass spectrometry methods were developed and validated to quantify sugars, sugar alcohols and amino acids. Specificity was achieved using multiple reaction monitoring, and the limits of quantification were in the pmole-fmole range. The comparative metabolomic study underlined that: (i) the majority of the metabolites accumulate in Physaria embryos between 18 and 27 DPA; (ii) the oxidative pentose phosphate pathway, glycolysis, the tricarboxilic acid cycle and the anaplerotic pathway drain a substantial amount of carbon; and (iii) ribulose-1,5-bisphosphate is present, which specifically indicates that the Calvin cycle is occurring. The importance and the relevance of these findings regarding fatty acid synthesis were discussed.

Biochemistry of hyperglycemia induced vascular dysfunction

Abstract Glucose enters the endothelium via a non-insulin sensitive GLUT-1 facilitated transporter that transports glucose continuously. Extracellular hyperglycemia is positively correlated with intracellular glucose. As the glucose levels increase, several changes in the glycolytic pathway, tricarboxilic acid cycle and the pentose phosphate pathway occur, all of them leading to an increase of oxygen reactive species (ROS). ROS are capable not only of directly impairing endothelial cells, but also indirectly by activating poly(ADPribose) polymerase, which inhibits glyceraldehyde 3-phosphate dehydrogenase. Further increase in glycloytic intermediates is followed by activation or overexpression of the main pathological pathways of hyperglycemia induced vascular damage: upregulation of glycation end-products, activation of protein kinase C, increased hexosamine pathway, and increased flux through polyol pathway, finally leading to the progressive narrowing and occlusion of the vessels.

Transcriptional and proteomic stress responses of a soil bacterium Bacillus cereus to nanosized zero-valent iron (nZVI) particles

Nanosized zero valent iron (nZVI) is emerging as an option for treating contaminated soil and groundwater even though the potentially toxic impact exerted by nZVI on soil microorganisms remains uncertain. In this work, we focus on nanotoxicological studies performed in vitro using commercial nZVI and one common soil bacterium (Bacillus cereus). Results showed a negative impact of nZVI on B. cereus growth capability, consistent with the entrance of cells in an early sporulation stage, observed by TEM. Despite no changes at the transcriptional level are detected in genes of particular relevance in cellular activity (narG, nirS, pykA, gyrA and katB), the proteomic approach used highlights differentially expressed proteins in B. cereus under nZVI exposure. We demonstrate that proteins involved in oxidative stress-response and tricarboxilic acid cycle (TCA) modulation are overexpressed; moreover proteins involved in motility and wall biosynthesis are repressed. Our results enable to detect a molecular-level response as early warning signal, providing new insight into first line defense response of a soil bacterium after nZVI exposure.

DIAGNOSIS OF ORGANIC ACIDEMIA

Organic acid occur as physiologic intermediates in variety of intracellular metabolic pathways, such as catabolism of aminoacid, mitochondrial β oxidation of fatty acids, tricarboxilic acid cycle, and cholestrol and fatty acid biosynthesis. The classical organic aciduria represent the pursuit of abnormalities of aminoacid degradation beyond deamination Their diagnostic hallmark is an accumulation of characteristic organic acids.The clinical features result from toxicity of the accumulating methabolites.Treatment involved 1. protein restriction 2. supplementation of aminoacids with unimpaired metabolism as well as trace elements and 3. specific measures for detoxification if indicated. Diagnostic tests consist of CBC, FBS, Bun, Cr, uric organic acid, TG, Cholestrol Ca, P, ALP, VBG, Na, K, Cl, U/A(PH, SG, Ketone), Ammonia, lactate, pyrovate, Ketone body CPK, Aldolase, SGOT, SGPT, BIL, PT, PTT, Plasma aminoacid HPLC, Homocysteine, Urine aminoacid and carbohydrate chromatography, Acyl carnitine profile, urine organic acids and for next steps tissue specimen and enzyme activity and gene study.clinical chemical indices of organid aciduria is Metabolic acidosis, Increased anion gap, Hyperglycemia and hypoglycemia, Ketosis and Ketonuria, Lactic acidosis, Hyperammonemia, Hyperuricemia, Hypertriglyceridemia, increase of transaminase Granulocytopenia, thrombocytopenia and Anemia. Acylcarnitine profile and urine organic acids are two for important tests for differentiation of types oforganic academia.

Taenia crassiceps: Host treatment alters glycolisis and tricarboxilic acid cycle in cysticerci

Human cysticercosis by Taenia crassiceps is rare although it is considered of zoonotic risk, especially to immunocompromised individuals. Albendazole and praziquantel are widely used and effective in its treatment. Their active forms inhibit the glucose uptake by the parasite and induce muscle contractions that alter its glycogen levels interfering in the energetic metabolism of the parasite and leading to its death. The aim of this study was to evaluate alterations in glycolysis, the tricarboxylic acid cycle and glucose concentrations caused by low dosage treatments of the hosts with albendazole and praziquantel. Therefore, T. crassiceps intraperitoneally infected mice were treated by gavage feeding with 5.75 or 11.5mg/kg of albendazole and 3.83 or 7.67mg/kg of praziquantel. The treated mice were euthanized after 24h and the cysticerci collected were morphologically classified into initial, larval or final phases. Concentrations of the organic acid produced and glucose were evaluated to detect alterations into the glycolysis and the tricarboxylic acid cycle pathways through chromatography and spectrophotometry. The low dosage treatment caused a partial blockage of the glucose uptake by the cysticerci in spite of the non significant difference between its concentrations. An activation of the tricarboxylic acid cycle was noted in the cysticerci that received the treatment due to an increase in the production of citrate, malate and α-ketoglutarate and the consumption of oxaloacetate, succinate and fumarate. The detection of α-ketoglutarate indicates that the cysticerci which were exposed to the drugs after host treatment present different metabolic pathways than the ones previously described after in vitro treatment.

Proposal of protocols using D-glutamine to optimize the 2,3-bis(2-methoxy-4-nitro-5-sulfophenly)-5-[(phenylamino) carbonyl]-2 H-tetrazolium hydroxide (XTT) assay for indirect estimation of microbial loads in biofilms of medical importance

Due to technical problems, biofilm biomasses are difficult to be precisely determined. One reliable strategy is based on the colorimetry of formazan compounds derived from tetrazolium salt reduction. XTT presents some desirable properties that make the biofilm measurements easier. However, cells entrapped within the extracellular matrixes normally do not metabolize the tetrazolium equally, leading to underestimation of cell contents. This study evaluated the effectiveness of d-glutamine, a plerotic substrate of tricarboxilic acid cycle (TAC), as inducer of XTT reduction. The metabolic activities of aerobic and anaerobic 48 h-old monospecific biofilms of Pseudomonas aeruginosa ATCC®27853™, Klebsiella pneumoniae ATCC®13883™, Staphylococcus epidermidis ATCC®12228™, Streptococcus mutans ATCC®25175™, and Candida albicans SC5314 were evaluated. Results showed that d-glutamine 50 mM (for P. aeruginosa, K. pneumoniae, and S. epidermidis) and 25 mM (for S. mutans and C. albicans) may enhance the detection of soluble formazan in a significant manner, what becomes the XTT reduction assay more robust.

Effect of low temperatures on glucose-induced insulin secretion and glucose metabolism in isolated pancreatic islets of the rat

The direct effect of hypothermia on the inhibition of insulin secretion may result from inhibition of the availability of energetic substrates and/or the lack of metabolic signals. In order to verify this hypothesis, the insulin secretion and the main metabolic glucose pathways were measured during the incubation of rat islets. In the presence of 16.7 mmol glucose/l and at 37 degrees C, insulin secretion was 925 +/- 119 microU/2 h per ten islets. With the same experimental conditions, glucose utilization, determined as the formation of 3H2O from [5-3H]glucose was 2225 +/- 184 pmol/2 h per ten islets, glucose oxidation measured as the formation of 14CO2 from [U-14C]glucose was 673 +/- 51 pmol/2 h per ten islets, pentose cycle determined as the formation of 14CO2 from either [1-14C]glucose or [6-14C]glucose was 37 +/- 5 pmol/2 h per ten islets; glucose oxidation by the tricarboxilic acid cycle, calculated to be the difference between glucose oxidation and pentose cycle values, was 636 pmol/2 h per ten islets. Hypothermia highly inhibited glucose-induced insulin secretion and glucose utilization. Inhibition of insulin secretion was partial at 27 degrees C since it was 2.5 times lower than that at 37 degrees C, and it was complete at 17 degrees C. Glucose oxidation in the tricarboxilic acid cycle was markedly inhibited by hypothermia since the inhibition coefficient (Q10) between 37 and 27 degrees C was 5. In contrast, glucose oxidation in the pentose phosphate shunt was enhanced at 27 degrees C, reaching 92 +/- 17 pmol/2 h per ten islets, and it was inhibited relatively little at 17 degrees C.(ABSTRACT TRUNCATED AT 250 WORDS)

Evaluation of the role of ionized calcium in sodium fluoroacetate (“1080”) poisoning

Fluoroacetate, which is an inhibitor of the tricarboxilic acid cycle, is widely used as a rodenticide. Fluoroacetate is converted in the body to fluorocitrate, which is an inhibitor of the enzyme aconitate hydrase. As a result, energy production goes down, and citrate accumulates. As citrate is a potent chelator of calcium ion, we postulated that ionized calcium concentration in the blood would drop. Fluoroacetate, 0.03 mmol/kg, was injected iv into anesthetized cats. Ionized calcium concentration in anaerobically drawn arterial blood samples was measured with an ion-exchange electrode. Samples were taken immediately before and 40 min after the poisoning, after which the animals were either used as controls (six cats) or treated with an infusion of iv CaCl 2 (another six cats), so as to restore ionized calcium levels to normal values. Forty minutes after fluoroacetate injection, the ionized calcium levels fell by an average of 27.2%, from 1.09 ± 0.07 to 0.79 ± 0.14 m M. There was a corresponding prolongation of the QTc interval of the ECG ( r = 0.82). Treatment with CaCl 2 significantly prolonged the life of the treated animals as compared to the control animals ( p < 0.0016 by the Mann-Whitney rank sum test). Our findings suggest that reduced levels of ionized calcium play an important role in the pathogenesis of fluoroacetate poisoning. The reduced levels of Ca 2+ are an adequate explanation for the toxic effects of fluoroacetate, and may be the missing link between the biochemistry of the poisoning and the clinical manifestations.

Mitochondria of Maize Weevil : Sitophilus zeamais (M.) : I. Isolation, Respiratory Function and Morphology of Adult Maize Weevil Mitochondria

Mitochondria isolated from the whole body of adult Sitophilus zeamais showed high rates of oxidation of pyruvate, α-glycerophosphate and various intermediates of the tricarboxilic acid cycle. The respiratory control ratio was 3.4-8.8 with pyruvate, 2.4-3.3 with α-ketoglutalate, 2.2-2.8 with malate, 1.8-2.3 with α-glycerophosphate, 1.8-2.2 with succinate and 1.3 with ascorbate+TMPD respectively as substrate. Respiration was accompanied by phosphorylation of ADP, with the expected ADP : O ratio for all substrates tested, indicating that there are three phosphorylation sites associated with the electron transport system. The maize weevil mitophodria showed a relatively high affinity for phosphate and RCR of the mitochodria was greatly influenced by Mg++. The purity and intactness of the mitochondria in morphology was also observed by electron microscopy.