RESUMO
The bacteria Mycobacterium tuberculosis is responsible for the infectious disease Tuberculosis. Targeting the tubercule bacteria is an important challenge in developing the antimycobacterials. The glyoxylate cycle is considered as a potential target for the development of anti-tuberculosis agents, due to its absence in the humans. Humans only possess tricarboxylic acid cycle, while this cycle gets connected to glyoxylate cycle in microbes. Glyoxylate cycle is essential to the Mycobacterium for its growth and survival. Due to this reason, it is considered as a potential therapeutic target for the development of anti-tuberculosis agents. Here, we explore the effect on the behavior of the tricarboxylic acid cycle, glyoxylate cycle and their integrated pathway with the bioenergetics of the Mycobacterium, under the inhibition of key glyoxylate cycle enzymes using Continuous Petri net. Continuous Petri net is a special Petri net used to perform the quantitative analysis of the networks. We first study the tricarboxylic acid cycle and glyoxylate cycle of the tubercule bacteria by simulating its Continuous Petri net model under different scenarios. Both the cycles are then integrated with the bioenergetics of the bacteria and the integrated pathway is again simulated under different conditions. The simulation graphs show the metabolic consequences of inhibiting the key glyoxylate cycle enzymes and adding the uncouplers on the individual as well as integrated pathway. The uncouplers that inhibit the synthesis of adenosine triphosphate, play an important role as anti-mycobacterials. The simulation study done here validates the proposed Continuous Petri net model as compared with the experimental outcomes and also explains the consequences of the enzyme inhibition on the biochemical reactions involved in the metabolic pathways of the mycobacterium.
Assuntos
Mycobacterium tuberculosis , Humanos , Metabolismo Energético , Ciclo do Ácido Cítrico/fisiologia , Antituberculosos/farmacologia , Antituberculosos/metabolismo , Glioxilatos/metabolismo , Glioxilatos/farmacologiaRESUMO
Monk fruit extract (MFE) is widely used as a sweetener in foods. In this study, the effects of the consumption of MFE-sweetened synbiotic yogurt on the lipid biomarkers and metabolism in the livers of type 2 diabetic rats were evaluated. The results revealed that the MFE-sweetened symbiotic yogurt affected the phosphatidylcholines, phosphatidylethanolamines, phosphatidylglycerol, lysophosphatidic acids, lysophosphatidylcholines, lysophosphatidylethanolamines, lysophosphatidylglycerols, lysophosphatidylinositols, lysophosphatidylserines, and fatty acid-hydroxy fatty acids biomarkers in the livers of type 2 diabetic rats. In addition, the consumption of the MFE-sweetened synbiotic yogurt significantly altered 12 hepatic metabolites, which are involved in phenylalanine metabolism, sphingolipid metabolism, bile secretion, and glyoxylate and dicarboxylate metabolism in the liver. Furthermore, a multiomics (metabolomic and transcriptomic) association study revealed that there was a significant correlation between the MFE-sweetened synbiotic yogurt and the metabolites and genes involved in fatty acid biosynthesis, bile secretion, and glyoxylate and dicarboxylate metabolism. The findings of this study will provide new insights on exploring the function of sweeteners for improving type 2 diabetes mellitus liver lipid biomarkers.
Assuntos
Cucurbitaceae , Diabetes Mellitus Experimental , Diabetes Mellitus Tipo 2 , Doenças dos Roedores , Simbióticos , Animais , Biomarcadores/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Diabetes Mellitus Tipo 2/veterinária , Ácidos Graxos/metabolismo , Frutas/química , Glioxilatos/metabolismo , Glioxilatos/farmacologia , Metabolismo dos Lipídeos , Lipídeos/farmacologia , Fígado/metabolismo , Extratos Vegetais/farmacologia , Ratos , Doenças dos Roedores/metabolismo , Edulcorantes/análise , Iogurte/análiseRESUMO
Multiple d-amino acids are present in mammalian cells, and these compounds have distinctive physiological functions. Among the free d-amino acids identified in mammals, d-aspartate plays critical roles in the neuroendocrine and endocrine systems, as well as in the central nervous system. Mammalian cells have the molecular apparatus necessary to take up, degrade, synthesize, and release d-aspartate. In particular, d-aspartate is degraded by d-aspartate oxidase (DDO), a peroxisome-localized enzyme that catalyzes the oxidative deamination of d-aspartate to generate oxaloacetate, hydrogen peroxide, and ammonia. However, little is known about the molecular mechanisms underlying d-aspartate homeostasis in cells. In this study, we established a cell line that overexpresses cytoplasm-localized DDO; this cell line cannot survive in the presence of high concentrations of d-aspartate, presumably because high levels of toxic hydrogen peroxide are produced by metabolism of abundant d-aspartate by DDO in the cytoplasm, where hydrogen peroxide cannot be removed due to the absence of catalase. Next, we transfected these cells with a complementary DNA library derived from the human brain and screened for clones that affected d-aspartate metabolism and improved cell survival, even when the cells were challenged with high concentrations of d-aspartate. The screen identified a clone of glyoxylate reductase/hydroxypyruvate reductase (GRHPR). Moreover, the GRHPR metabolites glyoxylate and hydroxypyruvate inhibited the enzymatic activity of DDO. Furthermore, we evaluated the effects of GRHPR and peroxisome-localized DDO on d- and l-aspartate levels in cultured mammalian cells. Our findings show that GRHPR contributes to the homeostasis of these amino acids in mammalian cells.
Assuntos
Oxirredutases do Álcool/metabolismo , Ácido Aspártico/metabolismo , Oxirredutases do Álcool/genética , Ácido Aspártico/farmacologia , Morte Celular/efeitos dos fármacos , Morte Celular/genética , Sobrevivência Celular/efeitos dos fármacos , D-Aspartato Oxidase/antagonistas & inibidores , D-Aspartato Oxidase/genética , D-Aspartato Oxidase/metabolismo , Glioxilatos/metabolismo , Glioxilatos/farmacologia , Células HEK293 , Células HeLa , Humanos , NADP , Piruvatos/metabolismo , Piruvatos/farmacologiaRESUMO
BACKGROUND AND PURPOSE: Urinary oxalate plays an important role in the formation of calcium oxalate renal stones, and approximately 50% to 60% of urinary oxalate is derived from the endogenous metabolism of glyoxylate. Therefore, we measured urinary oxalate, glycolate, glyoxylate, and citrate concentrations after acute intravenous administration of various doses of glyoxylate in rats to study oxalate metabolism. MATERIALS AND METHODS: Male Wistar rats weighing approximately 200 g were divided into six groups of eight animals each. Anesthetized rats received glyoxylate (0, 1, 2, 5, 10, and 20 mg) intravenously. Urine specimens were collected before and every hour after each dose for 4 hours, and the concentrations of oxalate, glycolate, glyoxylate, and citrate were measured by capillary electrophoresis. RESULTS: Hourly oxalate excretion in the urine peaked at 1 hour after glyoxylate administration, and the peak concentration increased in a dose-dependent manner. Approximately 15% to 30% (mol/mol) of the dose was converted to oxalate within 4 hours and 2% to 4.6% was converted to glycolate. Urinary glyoxylate was not detectable before glyoxylate administration, but large doses resulted in a significant amount of glyoxylate (0.7%-2.3%) appearing in the urine, and the level peaked at 1 hour after administration. Urinary glycolate also peaked at 1 hour after administration of glyoxylate. The urinary citrate concentration generally decreased by 3% to 33% after each dose of glyoxylate, except that it increased slightly after the 20-mg dose. CONCLUSION: Administration of glyoxylate increased urinary oxalate and glycolate excretion in rats, supporting the importance of the glycolate-glyoxylate-oxalate pathway.
Assuntos
Ácido Cítrico/urina , Glicolatos/urina , Glioxilatos/farmacologia , Glioxilatos/urina , Oxalatos/urina , Animais , Relação Dose-Resposta a Droga , Eletroforese Capilar , Glioxilatos/administração & dosagem , Injeções Intravenosas , Masculino , Ratos , Ratos Wistar , Fatores de TempoRESUMO
A potent lipid-lowering thyromimetic (CGS 26214) devoid of cardiac and thermogenic activity was identified based on its ability to preferentially access and bind the nuclear fraction of hepatocytes over that of myocytes in culture. The difference in access achieved with CGS 26214 was at least 100-fold better for hepatocytes than for myocytes. This in vitro hepatoselectivity resulted in a compound with unprecedented in vivo lipid-lowering potency with a minimal effective dose of 1 microgram/kg in rats and dogs (approximately 25x that of L-T3). At the same time, CGS 26214 was free of any cardiovascular effects up to the highest dose tested of 25 mg/kg and 100 micrograms/kg in rats and dogs, respectively.
Assuntos
Temperatura Corporal/efeitos dos fármacos , Sistema Cardiovascular/efeitos dos fármacos , Glioxilatos/farmacologia , Hipolipemiantes/farmacologia , Animais , Animais Recém-Nascidos , Anticolesterolemiantes/metabolismo , Anticolesterolemiantes/farmacologia , Ligação Competitiva , Carcinoma Hepatocelular/patologia , Débito Cardíaco/efeitos dos fármacos , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/metabolismo , Colesterol/sangue , Cães , Avaliação Pré-Clínica de Medicamentos , Glioxilatos/metabolismo , Coração/efeitos dos fármacos , Hipercolesterolemia/sangue , Hipolipemiantes/metabolismo , Fígado/efeitos dos fármacos , Neoplasias Hepáticas/patologia , Masculino , Contração Miocárdica/efeitos dos fármacos , Especificidade de Órgãos , Consumo de Oxigênio/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Receptores de LDL/metabolismo , Segurança , Hormônios Tireóideos/farmacologia , Triglicerídeos/sangue , Tri-Iodotironina/metabolismo , Células Tumorais CultivadasRESUMO
Parallel investigations of the transamination pathways of glutamine oxidation in Ehrlich ascites carcinoma (EAC) and AS 30D hepatoma revealed that hepatoma cells, unlike EAC, produce very little aspartate. This cannot be explained by differences in the activity of glutamine-metabolizing enzymes. Also, the mitochondria from the hepatoma respired at a similar rate to EAC mitochondria with glutamine as sole substrate producing substantial amounts of aspartate. Unlike their isolated mitochondria, intact hepatoma cells showed a very low rate of glutamine oxidation. Compared with EAC, the rate of L-[U-14C]glutamine consumption by AS 30D hepatoma cells was much lower, with insignificant production of 14C-labelled aspartate and CO2. This suggested that the glutamine-transporting system in the hepatoma cell plasma membrane had a very low activity. Isolated hepatoma mitochondria produced 3 times more pyruvate from malate than did EAC mitochondria, indicating a higher activity of NAD(P)-dependent malic enzyme. We postulate that an active malic enzyme may suppress the synthesis of aspartate in hepatoma cells, but further evidence is needed to confirm this assumption.
Assuntos
Glutaminase/metabolismo , Glutamina/metabolismo , Alanina/metabolismo , Animais , Ácido Aspártico/metabolismo , Radioisótopos de Carbono , Carcinoma de Ehrlich/enzimologia , Carcinoma de Ehrlich/metabolismo , Membrana Celular/enzimologia , Citratos/metabolismo , Ácido Cítrico , Citosol/enzimologia , Glioxilatos/farmacologia , Cinética , Neoplasias Hepáticas Experimentais/enzimologia , Neoplasias Hepáticas Experimentais/metabolismo , Camundongos , Mitocôndrias Hepáticas/efeitos dos fármacos , Mitocôndrias Hepáticas/enzimologia , Ratos , Ratos Endogâmicos , Especificidade por Substrato , Células Tumorais Cultivadas , UrânioRESUMO
The synthesis of hippurate from benzoate as compared to ureagenesis has been investigated in isolated rat hepatocytes. Half-maximal synthesis of hippurate was observed at 0.3 mmol/l benzoate. In the presence of 1 mmol/l benzoate, hippurate synthesis proceeded linearly with time at a rate of 40 +/- 10 mumol X h-1 X g-1 dry weight. This provided less than 10% of nitrogen epuration supported by concomitant urea synthesis (350 +/- 82 mumol X h-1 X g-1 dry weight). The incorporation of benzoate to hippurate was markedly limited by the availability of glycine. Half-maximal hippurate synthesis was observed at 2 mmol/l glycine. In the absence of glycine, piridoxilate, a glyoxylate derivative, markedly potentiated hippurate synthesis. Half-maximal stimulation was observed at 10 mmol/l piridoxilate. In the presence of 10 mmol/l piridoxilate, hippurate synthesis (420 +/- 35 mumol X h-1 X g-1 dry weight) provided more than 50% of nitrogen epuration supported by urea synthesis. It is concluded that supplementation with nitrogen-free analogues of glycine such as piridoxilate are required to potentiate hippurate synthesis in an attempt to replace ureagenesis as an alternative pathway of waste nitrogen excretion in inborn errors of urea synthesis.
Assuntos
Benzoatos/metabolismo , Glioxilatos/farmacologia , Hipuratos/biossíntese , Fígado/metabolismo , Piridoxina/análogos & derivados , Ureia/biossíntese , Alanina/metabolismo , Erros Inatos do Metabolismo dos Aminoácidos/metabolismo , Animais , Células Cultivadas , Relação Dose-Resposta a Droga , Glicina/farmacologia , Hipuratos/urina , Masculino , Ornitina/metabolismo , Piridoxina/farmacologia , Ratos , Ratos Endogâmicos , Estimulação QuímicaRESUMO
Urinary endogenous oxalate was increased by feeding vitamin B6-deficient or control rats with 5.2% hydroxyproline, or 3% glycine plus 5.2% hydroxyproline. The activities of liver lactic dehydrogenase (LDH), glucose-6-phosphate dehydrogenase (G6PD) , malic enzyme (ME), and ATP citrate lyase were decreased in vitamin B6-DEFICIENT RATS, AND THEIR LIVEr G6PD was further decreased by the addition of glycine and hydroxyproline to their diets. Supplementing control diets with the two amino acids decreased the activities of rat liver LDH, G6PD, and ATP citrate lyase. The effects of glycine and hydroxyproline feeding on the enzymes studied did not appear related to alterations in insulin availability. Since in vitamin B6-deficient rats, there are increases in urinary levels of oxalic and glycolic acids, and glycine, and increases in tissue levels of glyoxylic acid and glycine, the effects of these metabolites on the activities of the above mentioned enzymes were measured. Oxalic acid inhibited the activities of LDH, G6PD, and ME. Glyoxylic acid inhibited LDH and ME, but not G6PD. Glycolic acid inhibited G6PD and ME, but not LDH. ATP citrate lyase was not affected by these substances. Glycine had no effect on the enzymes studied. Diets which increased oxalate excretion generally reduced or did not alter liver and kidney levels of oxalate, glycolate, and glyoxylate. However, the feeding of glycine and hydroxyproline increased kidney oxalate, and liver and kidney glyoxylate in vitamin B6-deficient rats.