Evaluation of Serum and Urine Amino Acids in Dogs with Chronic Kidney Disease and Healthy Dogs Fed a Renal Diet

This observational study aimed to evaluate serum and urinary amino acid (AA) concentrations in healthy dogs and dogs with chronic kidney disease (CKD) fed a commercial therapeutic renal diet with reduced protein and phosphorus levels. Ten dogs with CKD stages 3 or 4 composed the study group and received the renal diet for 180 days (RG T180). A control group (CG T30) composed of seven healthy dogs was fed a renal diet for 30 days. When comparing serum AA between RG T180 and CG T30, histidine, isoleucine, leucine, lysine, phenylalanine, tryptophan, cysteine, citrulline, ornithine, taurine, branched-chain amino acids (BCAA), and total essential amino acids (EAA) were higher in RG T180. Meanwhile, arginine, asparagine, aspartate, glutamine, serine, and tyrosine were higher in CG T30. Serum phenylalanine, tryptophan, and hydroxyproline were higher in RG T0 (dogs with CKD before consuming a renal diet) when compared to RG T180. In addition, the serum ratios of arginine/citrulline, tyrosine/phenylalanine, and serine/glycine were higher in CG T30 than in RG T180. Concerning urinary AA concentrations in CKD dogs, isoleucine, phenylalanine, tryptophan, aspartate, cysteine, and BCAA were higher in RG T180. In urine, the total EAA/total non-essential AA ratio in RG T180 was higher than in CG T30 as well as tyrosine/phenylalanine ratio higher in CG T30. In conclusion, the combination of renal diet and conservative treatment over 6 months in dogs with CKD stages 3 or 4 affected the AAs metabolism when compared to healthy adult dogs.

Abstract P165: Urinary Amino Acid Profile And Cardiovascular Parameters In Overweight And Obese Brazilian Children

Despite being thoroughly studied, there isn’t complete elucidation of mechanisms underlying obesity, and especially regarding overweight in children. Small molecule metabolites are promising candidates for understanding obesity′s phenotypes. Recent metabolic studies have advanced in discovery of biomarkers for better knowledge of mechanisms related to obesity and its comorbidities. In this cross-sectional study it was evaluated the profile of urinary amino acids of 110 children that were classified in into groups of nutritional status according to their BMI/A (Body Mass Index/age), as normal weight (NW) (n=45), overweight (OW) (n=21) and obese (OB) (n=44). The 24h urine samples were submitted to LC-MS/MS for evaluation of 47 amino acids using Amino Acids Analysis Kit (Zivak®, Turkey), corrected by creatinine concentration. Biochemical profile, cardiovascular parameters and anthropometric measurements were assessed following standard protocols. Girls showed a higher heart rate (G: 82bpm vs B:79bpm, p=0,01 ) than boys. Increased weight presented higher values of systolic blood pressure ( NW: 102.0, OW: 109.0, OB: 110.0; NW vs OW, p = 0.0001; NW vs OB, p = 0.0001 ) and diastolic blood pressure ( NW: 60.0, OW: 67.0, OB: 66.75, NW vs OW, p < 0.0001; NW vs OB, p < 0.0001 ). The following amino acids (expressed in nmol/mg of creatinine) were found in higher concentrations in children’s urine with overweight or obesity, when compared with NW group, glutamine ( NW: 1024.3 vs OW: 1309.2, p=0.05), tryptophan (NW: 85.65, OW:113.55, OB:109.36; NW vs OW p=0.01; OW vs OB p=0.004), phenylalanine (NW: 43.04, OW:49.66, OB:72.08; NW vs OB p=0.01; OW vs OB p=0.04), and Tyrosine (NW: 85.07, OW: 109.74, OB: 140.54; NW vs OB p<0.01; OW vs OB p=0.04). In contrast, sarcosine had reduced concentrations for groups with weight excess (OW: 78.7 vs OB: 62.3, p=0.04). Urine levels of glutamine and aromatic amino acids were higher in OW and OB groups. Tyrosine also showed a positive association with systolic blood pressure values. Thus, there was a positive correlation between levels of glutamine and aromatics amino acids and excess weight, suggesting that they may be candidates to biomarkers for obesity in children. Furthermore, obesity was directly related to increase in blood pressure values.

Profile of urinary amino acids and their post-translational modifications (PTM) including advanced glycation end-products (AGEs) of lysine, arginine and cysteine in lean and obese ZSF1 rats

Heart failure with preserved ejection fraction (HFpEF) is associated with high mortality and has an increasing prevalence associated with the demographic change and limited therapeutic options. Underlying mechanisms are largely elusive and need to be explored to identify specific biomarkers and new targets, which mirror disease progression and intervention success. Obese ZSF1 (O-ZSF1) rats are a useful animal model, as they spontaneously develop hypertension, hyperlipidemia and glucose intolerance and finally HFpEF. The urinary profile of amino acids and their metabolites of post-translational modifications (PTM), including the advanced glycation end-products (AGEs) of lysine, arginine and cysteine, are poorly investigated in HFpEF and ZSF1 rats. The aim of the present study was to characterize the status of free amino acids and their metabolites of PTM and glycation in lean ZSF1 (L-ZSF1) and O-ZSF1 rats in urine aiming to find possible effects of glucose on the excretion of native and modified amino acids. In the urine of twelve L-ZSF1 and twelve O-ZFS1 rats collected at the age of 20 weeks, we measured the concentration of native and modified amino acids by reliable previously validated stable-isotope dilution gas chromatography-mass spectrometry (GC–MS) approaches. Serum glucose was 1.39-fold higher in the O-ZSF1 rats, while urinary creatinine concentration was 2.5-fold lower in the O-ZSF1 rats. We observed many differences in urinary amino acids excretion between L-ZSF1 and O-ZSF1 rats. The creatinine-corrected homoarginine excretion was twofold lower in the O-ZSF1 rats. We also observed distinct associations between the concentrations of serum glucose and urinary amino acids including their PTM and AGE metabolites in the L-ZSF1 and O-ZSF1 rats. Our study shows that PTM metabolites and AGEs are consistently lower in the L-ZSF1 than in the O-ZSF1 rats. Serum malondialdehyde (MDA) concentration was higher in the O-ZSF1 rats. These results suggest that hyperglycemia, hyperlipidemia and elevated oxidative stress in the O-ZSF1 rats favor PTM methylation of arginine and lysine and the glycation of lysine and cysteine. The area under the receiver operation characteristic (ROC) curve values were 0.996 for serum glucose, 0.951 for urinary creatinine, 0.939 for serum MDA, 0.885 for Nε-carboxyethyl-lysine, 0.830 for carboxyethyl-cysteine, and 0.792 for monomethyl-lysine. Non-invasive measurement of methylation and glycation products of arginine, lysine and cysteine residues in proteins in urine of L-ZSF1 and O-ZSF1 rats may be useful in studying pathophysiology and pharmacology of HFpEF.

Diagnostic biomarkers for USD via serum and urinary amino acid profiles using a metabolomics-based strategy

Background: Urinary stone disease (USD) is a common disease with a high prevalence, a high recurrence rate and a lack of effective diagnostic biomarkers.Methods: To explore the potential biomarkers for USD early diagnosis, the serum and urinary amino acid (AA) profiles of 74 USD patients were compared with 35 healthy controls (HC) via gas chromatography coupled with mass spectrometry (GC-MS).Results: Based on the serum and urinary AA profiles analyzed by metabolomic strategy, a group of 5 differentially expressed AAs including Ser, Glu, Asp, Ile and Gly and 7 metabolic pathway disorders in patients were revealed.Conclusions: These serum and urinary AA profiles could provided clinical biomarkers for USD early diagnosis, and also indicated that some specific AA metabolic pathways might play important roles in the pathogenesis of USD.

Plasma and Urinary Amino Acid-Derived Catabolites as Potential Biomarkers of Protein and Amino Acid Deficiency in Rats

Objective: Dietary intakes must cover protein and essential amino acid (EAA) requirements. For this purpose, different methods have been developed such as the nitrogen balance method, factorial method, or AA tracer studies. However, these methods are either invasive or imprecise, and the Food and Agriculture Organization of the United Nations (FAO, 2013) recommends new methods and, in particular, metabolomics. The aim of this study is to determine total protein/EAA requirement in the plasma and urine of growing rats. Methods: 36 weanling rats were fed with diets containing 3, 5, 8, 12, 15, and 20% protein for 3 weeks. During experimentation, urine was collected using metabolic cages, and blood from the portal vein and vena was taken at the end of the experiment. Metabolomics analyses were performed using LC-MS, and the data were analyzed with a multivariate analysis model, partial least Squares (PLS) regression, and independent component-discriminant analysis (ICDA). Each discriminant metabolite identified by PLS or ICDA was tested by one-way ANOVA to evaluate the effect of diet. Results: PLS and ICDA allowed us to identify discriminating metabolites between different diet groups. Protein deficiency led to an increase in the AA catabolism enzyme systems inducing the production of breakdown metabolites in the plasma and urine. Conclusion: These results indicate that metabolites are specific for the state of EAA deficiency and sufficiency. Some types of biomarkers such as AA degradation metabolites appear to be specific candidates for protein/EAA requirement.

Diagnostic Approach in Cystinuria: A Case Report

Cystinuria is an, inherited metabolic disorder progressing with recurrent kidney stones due to impaired reabsorption of dibasic amino acids and arises from mutations in the SLC3A1 and SLC7A9 on chromosome 2. Cystine crystals were detected in the urinalysis of a 17-year-old male patient who was investigated for recurrent kidney stones. Because of demonstration of cystine excretion in the urinary amino acid analysis and having positive family history, we suspected Cystinuria Type B and initiated supportive therapy. However, based on the results of molecular analyses his diagnosis was changed as Cystinuria Type A. In conclusion, our final diagnosis was changed according to the molecular analyses but our treatment approach did not change. Therefore we would like to emphasize that, prominent physical examination findings and supportive laboratory test results will be sufficient for the diagnosis of cystinuria.

Effects of Aging, Long-Term and Lifelong Exercise on the Urinary Metabolic Footprint of Rats

Life expectancy has risen in the past decades, resulting in an increase in the number of aged individuals. Exercise remains one of the most cost-effective treatments against disease and the physical consequences of aging. The purpose of this research was to investigate the effects of aging, long-term and lifelong exercise on the rat urinary metabolome. Thirty-six male Wistar rats were divided into four equal groups: exercise from 3 to 12 months of age (A), lifelong exercise from 3 to 21 months of age (B), no exercise (C), and exercise from 12 to 21 months of age (D). Exercise consisted in swimming for 20 min/day, 5 days/week. Urine samples collection was performed at 3, 12 and 21 months of life and their analysis was conducted by liquid chromatography-mass spectrometry. Multivariate analysis of the metabolite data did not show any discrimination between groups at any of the three aforementioned ages. However, multivariate analysis discriminated the three ages clearly when the groups were treated as one. Univariate analysis showed that training increased the levels of urinary amino acids and possibly protected against sarcopenia, as evidenced by the higher levels of creatine in the exercising groups. Aging was accompanied by decreased levels of urinary amino acids and signs of increased glycolysis. Concluding, both aging and, to a lesser degree, exercise affected the rat urinary metabolome, including metabolites related to energy metabolism, with exercise showing a potential to mitigate the consequences of aging.