Urea Cycle Metabolites Research Articles

Abstract 12306: Changes of Metabolomic Profiling Are Associated With Reduced Exercise Capacity in Patients With Heart Failure

Background: Exercise intolerance is a major clinical manifestation, and closely related to poor prognosis in heart failure (HF) patients. Metabolic derangements have been shown to play an important role in this phenomenon. Recent metabolomic investigations have highlighted several novel biochemical pathways in HF, however, metabolic profiling associated with exercise capacity has not been elucidated. We thus investigated the metabolites associated with exercise capacity by using global metabolomic analysis of plasma samples in HF patients. Methods: Ten consecutive HF patients with reduced ejection fraction (57 ± 12 years old, New York Heart Association class II - III) and 5 age-matched healthy subjects as controls (56 ± 3 years old) were studied. Charged metabolites in plasma samples were measured by capillary electrophoresis mass spectrometry. Peak oxygen uptake (peak VO 2 ) and VE/VCO 2 slope were measured by cardiopulmonary exercise test. Results: A total of 233 charged metabolites were identified and quantified absolutely using standard chemicals for each metabolite. Significant increases in arginine methylation, phospholipid metabolites, and urea cycle metabolites were observed in HF patients compared to controls (Figure). Peak VO 2 was negatively correlated with methylarginine including asymmetric dimethylarginine (ADMA) (r=-0.77, P<0.001) and symmetric dimethylarginine (SDMA) (r=-0.65, P=0.040) levels, and phospholipid metabolites including choline (r=-0.63, P=0.049) and N,N -dimethylglycine (r=-0.75, P=0.011) levels in HF patients. VE/VCO 2 slope was positively correlated with N,N -dimethylglycine (r=0.72, P=0.026) and Urea (r=0.82, P=0.006) levels. Conclusions: Increases in metabolites of arginine methylation, phospholipid metabolism, and urea cycle were associated with exercise intolerance in HF. These results provide mechanistic insights into exercise intolerance associated with HF.

Metabolite Profiles During Oral Glucose Challenge

To identify distinct biological pathways of glucose metabolism, we conducted a systematic evaluation of biochemical changes after an oral glucose tolerance test (OGTT) in a community-based population. Metabolic profiling was performed on 377 nondiabetic Framingham Offspring cohort participants (mean age 57 years, 42% women, BMI 30 kg/m2) before and after OGTT. Changes in metabolite levels were evaluated with paired Student t tests, cluster-based analyses, and multivariable linear regression to examine differences associated with insulin resistance. Of 110 metabolites tested, 91 significantly changed with OGTT (P ≤ 0.0005 for all). Amino acids, β-hydroxybutyrate, and tricarboxylic acid cycle intermediates decreased after OGTT, and glycolysis products increased, consistent with physiological insulin actions. Other pathways affected by OGTT included decreases in serotonin derivatives, urea cycle metabolites, and B vitamins. We also observed an increase in conjugated, and a decrease in unconjugated, bile acids. Changes in β-hydroxybutyrate, isoleucine, lactate, and pyridoxate were blunted in those with insulin resistance. Our findings demonstrate changes in 91 metabolites representing distinct biological pathways that are perturbed in response to an OGTT. We also identify metabolite responses that distinguish individuals with and without insulin resistance. These findings suggest that unique metabolic phenotypes can be unmasked by OGTT in the prediabetic state.

Integrated profiling of metabolites and trace elements reveals a multifaceted malnutrition in pregnant women from a region with a high prevalence of congenital malformations

Considerable evidence from epidemiological and clinical studies demonstrated that maternal nutritional status is closely associated with placental, embryonic, fetal growth and development, and ultimately pregnancy outcomes. In recent years, there has been a growing interest in the study of disorders of pregnancy using a metabolomic profiling approach. In this study, we presented an integrated comprehensive profiling approach to assess maternal nutritional status through measuring a wide variety of small-molecule metabolites and trace elements in serum of pregnant women. A total of 56 pregnant women with normal pregnancy outcomes were enrolled from Lvliang prefecture of Shanxi province, the area with the highest prevalence of congenital anomalies in China, and 40 pregnant women with normal pregnancies were recruited from Huairou county of Beijing city, the region representing a national average level. As compared with the national average level, these pregnant women from Lvliang region shown distinct metabolic phenotypic variations as revealed by the depleted serum concentrations of folate and vitamin B12, lower concentrations of carbohydrates, lipids, Se, Zn, and Cu, as well as higher concentrations of amino acids, urea-cycle metabolites, Sr, Cd, and Pb. Our results offer an improved understanding of severe multifaceted malnutrition in the pregnant women from a population with a high prevalence of congenital anomalies, highlighting the potential of a panel of critical nutrients as markers for aiding the diagnosis, prevention, and intervention of pregnancy complications.

Toward Metabolomic Signatures of Cardiovascular Disease

Small biochemicals are the end result of all the regulatory complexity present in a cell, tissue, or organism, including transcriptional regulation, translational regulation, and posttranslational modifications. Metabolic changes are thus the most proximal reporters of the body’s response to a disease process or drug therapy. In 1971, Robinson and coworkers1 conceived the core idea that information-rich data reflecting the functional status of a complex biological system resides in the quantitative and qualitative pattern of metabolites in body fluids. In the same year, Horning and Horning2 first used the term metabolic profiling to describe the output of a gas chromatogram from a patient sample. This new approach to the quantitative metabolic profiling of large numbers of small molecules in biofluids was ultimately termed “metabonomics” by Nicholson et al3 and “metabolomics” by others. Article see p 207 Two core technologies are used to perform metabolic profiling: nuclear magnetic resonance and tandem mass spectrometry (MS/MS), as previously reviewed in Circulation: Cardiovascular Genetics. 4 Nuclear magnetic resonance requires relatively little sample preparation and is nondestructive, allowing for subsequent structural analyses. However, the method tends to have low sensitivity and can detect only highly abundant analytes. Tandem mass spectrometry (MS/MS), coupled with liquid chromatography, on the other hand, has much higher sensitivity for small molecules and is also applicable to a wide range of biological fluids (including serum, plasma, and urine). Recent advances in MS technology now enable researchers to determine analyte masses with such high precision and accuracy that metabolites can be identified unambiguously even in complex fluids. These technologies can be used to characterize biological samples either in a targeted manner or in a pattern discovery manner. In the former, the investigator targets a predefined …

SIRT3 Promotes the Urea Cycle by Deacetylating Ornithine Transcarbamoylase

Identifying the substrates for SIRT3 is paramount to understanding how reversible acetylation can modulate mitochondrial metabolism. Here we utilize a SPOT peptide library containing ~250 unique 9‐mer peptide sequences of previously identified acetylated mitochondrial proteins. Within the nine residue peptides, a central thiotrifluoro‐acetyl‐lysine was used as a tight‐binding mimic of acetyl‐lysine. Library screening revealed a wide range in binding affinity, which was validated by measuring the catalytic efficiency of deacetylation of the corresponding free peptides. One particular peptide that conferred very strong binding affinity was lysine 88 of ornithine transcarbamoylase(OTC) previously identified to be regulated by acetylation. OTC converts ornithine and carbamoyl phosphate into citrulline, and is a key enzyme in urea cycle to detoxify ammonium produced from amino acid metabolism. Reversible acetylation of lysine 88 plays an important role in regulating OTC activity through responding to cellular metabolic status. SIRT3 interacts with and deacetylates OTC on lysine 88, upregulating its activity. Knockdown of SIRT3 in cultured cells elevates OTC acetylation decreasing activity. Utilizing SIRT3−/− mice, OTC displays increased acetylation and decreased activity. Furthermore, measurements of urea cycle metabolites reveal defects in the SIRT3−/− animals. These findings demonstrate that SIRT3 regulates OTC and the ability to adapt to low energy input, avoiding a toxic buildup of ammonium from amino acid catabolism.

Association of a Peripheral Blood Metabolic Profile With Coronary Artery Disease and Risk of Subsequent Cardiovascular Events

Molecular tools may provide insight into cardiovascular risk. We assessed whether metabolites discriminate coronary artery disease (CAD) and predict risk of cardiovascular events. We performed mass-spectrometry-based profiling of 69 metabolites in subjects from the CATHGEN biorepository. To evaluate discriminative capabilities of metabolites for CAD, 2 groups were profiled: 174 CAD cases and 174 sex/race-matched controls ("initial"), and 140 CAD cases and 140 controls ("replication"). To evaluate the capability of metabolites to predict cardiovascular events, cases were combined ("event" group); of these, 74 experienced death/myocardial infarction during follow-up. A third independent group was profiled ("event-replication" group; n=63 cases with cardiovascular events, 66 controls). Analysis included principal-components analysis, linear regression, and Cox proportional hazards. Two principal components analysis-derived factors were associated with CAD: 1 comprising branched-chain amino acid metabolites (factor 4, initial P=0.002, replication P=0.01), and 1 comprising urea cycle metabolites (factor 9, initial P=0.0004, replication P=0.01). In multivariable regression, these factors were independently associated with CAD in initial (factor 4, odds ratio [OR], 1.36; 95% CI, 1.06 to 1.74; P=0.02; factor 9, OR, 0.67; 95% CI, 0.52 to 0.87; P=0.003) and replication (factor 4, OR, 1.43; 95% CI, 1.07 to 1.91; P=0.02; factor 9, OR, 0.66; 95% CI, 0.48 to 0.91; P=0.01) groups. A factor composed of dicarboxylacylcarnitines predicted death/myocardial infarction (event group hazard ratio 2.17; 95% CI, 1.23 to 3.84; P=0.007) and was associated with cardiovascular events in the event-replication group (OR, 1.52; 95% CI, 1.08 to 2.14; P=0.01). Metabolite profiles are associated with CAD and subsequent cardiovascular events.

Metabolite profiling of human colon carcinoma – deregulation of TCA cycle and amino acid turnover

BackgroundApart from genetic alterations, development and progression of colorectal cancer has been linked to influences from nutritional intake, hyperalimentation, and cellular metabolic changes that may be the basis for new diagnostic and therapeutic approaches. However, in contrast to genomics and proteomics, comprehensive metabolomic investigations of alterations in malignant tumors have rarely been conducted.ResultsIn this study we investigated a set of paired samples of normal colon tissue and colorectal cancer tissue with gas-chromatography time-of-flight mass-spectrometry, which resulted in robust detection of a total of 206 metabolites. Metabolic phenotypes of colon cancer and normal tissues were different at a Bonferroni corrected significance level of p = 0.00170 and p = 0.00005 for the first two components of an unsupervised PCA analysis. Subsequent supervised analysis found 82 metabolites to be significantly different at p < 0.01. Metabolites were connected to abnormalities in metabolic pathways by a new approach that calculates the distance of each pair of metabolites in the KEGG database interaction lattice. Intermediates of the TCA cycle and lipids were found down-regulated in cancer, whereas urea cycle metabolites, purines, pyrimidines and amino acids were generally found at higher levels compared to normal colon mucosa.ConclusionThis study demonstrates that metabolic profiling facilitates biochemical phenotyping of normal and neoplastic colon tissue at high significance levels and points to GC-TOF-based metabolomics as a new method for molecular pathology investigations.

ARGININE CHALLENGE UNRAVELS PERSISTENT DISTURBANCES OF UREA CYCLE AND GLUCONEOGENESIS IN ABSTINENT ALCOHOLICS

Data on recovery from hormonal and metabolic sequelae of alcoholism in strictly controlled alcohol abstinence are mainly restricted to short-term abstention. Our previous findings of persistently decreased plasma and urinary urea concentrations in long-term abstinent alcoholics prompted us to further elucidate this unexplained phenomenon. The response of circulating urea cycle metabolites and glucose-regulating hormones to an intravenous load (30 g) of arginine hydrochloride was investigated in abstinent male alcoholics (n = 14) after complete recovery of all routine liver parameters and compared with that in healthy male controls (n = 15). The arginine challenge provoked (i) higher peak concentrations of arginine and increased arginine/ornithine and ornithine/citrulline ratios in the plasma of abstinent alcoholics; (ii) augmented plasma glutamine concentrations in alcoholics in the presence of comparable levels in both experimental groups of plasma glutamate, ammonia, and nitrate/nitrite; (iii) parallel increases in plasma urea concentrations over the respective baseline levels but distinctly higher urinary urea excretion in controls; (iv) a blunted blood glucose response to arginine in alcoholics together with a reduced insulin and glucagon surge; and (v) an elevated growth hormone peak as compared with controls. Application of an intravenous arginine challenge reveals profound and lasting metabolic and hormonal disturbances in abstinent alcoholics, affecting urea cycle and gluconeogenesis. The common denominator of many of these changes may be an acquired irreversible deficiency in cellular energy regulation.

Dietary and metabolic manipulations of the carcinogenic process: role of nucleotide pool imbalances in carcinogenesis.

Perturbations in DNA and/or membranes are considered to be important for the carcinogenic process. A search for nutritional and metabolic means of disturbing the homeostasis of DNA and membranes revealed that nucleotide pools offer an exciting possibility. An imbalance in nucleotide pools can exert a two-pronged attack on both DNA and membranes. When given to rats, orotic acid, a precursor of pyrimidine nucleotides, results in an imbalance in nucleotide pools (an increase in uridine nucleotides and a decrease in inosine/adenine nucleotides), alterations in both DNA and membranes, and promotion of carcinogenesis in the liver initiated by chemical carcinogens. Agents such as adenine and allopurinol, which inhibit the metabolism of orotic acid and thereby decrease the formation of uridine nucleotides, and galactosamine, which traps uridine nucleotides, inhibited the promotional effects of orotic acid in the liver. These results suggested that orotic acid needs to be metabolized to uridine nucleotides and the creation of a subsequent imbalance in nucleotide pools is important for the promotional effects of orotic acid. To determine whether the creation of a nucleotide pool imbalance is a more general mechanism of tumor promotion, two lines of approach were investigated. One was to determine the effect of orotic acid on promotion of carcinogenesis in other organs, and the second approach was to determine how to induce nucleotide pool imbalances by means other than orotic acid administration. It is interesting to note that orotic acid promotes carcinogenesis in duodenum initiated by azoxymethane. Regarding the second approach, it became apparent that several metabolic disturbances result in increased orotic acid synthesis and alterations in nucleotide pools.(ABSTRACT TRUNCATED AT 250 WORDS)

DIETARY EFFECTS ON THE CONCENTRATIONS OF FREE AMINO ACIDS IN PLASMA AND WHOLE BLOOD OF PIGS

The concentrations of free amino acids in plasma and whole blood of growing pigs were examined following an overnight fast and at 2 and 5 h after eating either a semipurified diet containing casein or a natural grain-based commercial ration. The plasma:whole blood (P:B) ratio of the concentrations of threonine, proline and tyrosine changed significantly (P &lt; 0.01) with time after feeding; methionine and lysine showed the same effect depending upon which diet was fed. Aspartic acid, citrulline, ornithine and histidine tended to show similar responses. No effect was observed on the ratios for other amino acids. Treatment effects on plasma:erythrocyte ratios were similar to those on P:B ratios. The semipurified diet resulted in increasing plasma and blood concentrations of most amino acids for 5 h after feeding whereas concentrations of most amino acids decreased after 2 h when the natural diet was fed. The natural diet appeared to stimulate greater urea cycle activity according to plasma concentrations of urea cycle metabolites. The data indicate that for some amino acids metabolic state affects plasma and whole blood concentrations differently, especially when a semipurified diet is fed. Whole blood concentrations of these amino acids are not an accurate reflection of plasma concentrations.