Quantitative amino acid and proteomic analysis: Very low excretion of polypeptides &gt;750 Da in normal urine

Nitric oxide (NO) production is diminished in many patients with cardiovascular and renal disease. Asymmetric dimethylarginine (ADMA) is an endogenous inhibitor of NO synthesis, and elevated plasma levels of ADMA are associated with poor outcomes. Dimethylarginine dimethylaminohydrolase-1 (DDAH1) is a methylargininemetabolizing enzyme that reduces ADMA levels. We reported previously that a DDAH1 gene variant associated with increased renal DDAH1 mRNA transcription and lower plasma ADMA levels, but counterintuitively, a steeper rate of renal function decline. Here, we test the hypothesis that reduced renal-specific ADMA metabolism protects against progressive renal damage. Renal DDAH1 is expressed predominately within the proximal tubule. A novel proximal tubule-specific Ddah1 knockout (Ddah1) mouse demonstrated tubular cell accumulation of ADMA and lower NO concentrations, but unaltered plasma ADMA concentrations. Ddah1 PT2/2 mice were protected from reduced kidney tissue mass, collagen deposition, and profibrotic cytokine expression in two independent renal injury models: folate nephropathy and unilateral ureteric obstruction. Furthermore, a study of two independent kidney transplant cohorts revealed higher levels of human renal allograft methylargininemetabolizing enzyme gene expression associated with steeper function decline. We also report an association among DDAH1 expression, NO activity, and uromodulin expression supported by data from both animal and human studies, raising the possibility that kidney DDAH1 expression exacerbates renal injury through uromodulinrelated mechanisms. Together, these data demonstrate that reduced renal tubular ADMA metabolism protects against progressive kidney function decline. Thus, circulating ADMA may be an imprecise marker of renal methylarginine metabolism, and therapeutic ADMA reduction may even be deleterious to kidney function.

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“…51 All values corrected for creatinine concentration. Mann-Whitney U test, n=8 (Ddah1 fl/fl ) and n=20 (Ddah1 PT2/2 ).…”

Section: Effect Of Pt-specific Ddah1 Deletion In Folate Nephropathymentioning

confidence: 99%

Reduced Renal Methylarginine Metabolism Protects against Progressive Kidney Damage

Tomlinson

Caplin

Boruc

et al. 2015

Journal of the American Society of Nephrology

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“…Urine is enriched relative to plasma in low molecular weight peptide components and amino acid derivatives such as hippuric acid and phenylacetylglutamine (47,70,71 ). On a molar basis, the urinary concentrations of these compounds usually exceed that of albumin, so there may be substantial ligand binding to albumin (70,71 ).…”

Section: Albumin Structurementioning

confidence: 99%

Current Issues in Measurement and Reporting of Urinary Albumin Excretion

et al. 2009

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BACKGROUND: Urinary excretion of albumin indicates kidney damage and is recognized as a risk factor for progression of kidney disease and cardiovascular disease. The role of urinary albumin measurements has focused attention on the clinical need for accurate and clearly reported results. The National Kidney Disease Education Program and the IFCC convened a conference to assess the current state of preanalytical, analytical, and postanalytical issues affecting urine albumin measurements and to identify areas needing improvement. CONTENT:The chemistry of albumin in urine is incompletely understood. Current guidelines recommend the use of the albumin/creatinine ratio (ACR) as a surrogate for the error-prone collection of timed urine samples. Although ACR results are affected by patient preparation and time of day of sample collection, neither is standardized. Considerable intermethod differences have been reported for both albumin and creatinine measurement, but trueness is unknown because there are no reference measurement procedures for albumin and no reference materials for either analyte in urine. The recommended reference intervals for the ACR do not take into account the large intergroup differences in creatinine excretion (e.g., related to differences in age, sex, and ethnicity) nor the continuous increase in risk related to albumin excretion. DISCUSSION:Clinical needs have been identified for standardization of (a) urine collection methods, (b) urine albumin and creatinine measurements based on a complete reference system, (c) reporting of test results, and (d) reference intervals for the ACR.

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“…According to the requirements of the producer the samples to be analyzed by the MALDI-TOF mass spectrometry must be filtered to separate ionic compounds, heavy metals, compounds of mass < 1 kDa from the protein (Saito et al 2010, Cobzac andGocan 2011). "Light" fraction can be qualitatively analyzed thanks to a special procedure (Norden et al 2004) and modification of detection method which is impossible in the case of 2D electrophoresis. Owing to the use of a precolumn of the SCX type in the Nano-LC system it is possible to separate over 2.5 thousand compounds from mixture.…”

Section: Liquid Chromatographymentioning

confidence: 99%

Applicability of 2D gel electrophoresis and liquid chromatography in proteomic analysis of urine using mass spectrometry MALDI-TOF

Banach¹,

Adaszek²,

Wyłupek³

et al. 2013

Polish Journal of Veterinary Sciences

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Proteomics including the studies of the structure, function and dependences between proteins is more and more extensively applied in human medicine and veterinary medicine. The analysis of protein profiles of tissues and body fluid from healthy and ill individuals allows to identify diagnostic, prognostic and predictive markers in various pathological states in people and animals. This paper presents preparation of urine samples for analysis in the mass spectrometer MALDI-TOF (Ultraflextreme, Bruker, Bremen, Germany) by means of two methods: liquid chromatography based on the system Nano-LC (PROTEINER FC II, Bruker Daltonics, Bremen Germany). and two-direction electrophoresis 2DE (GE Healthcare, United Kingdom). Both methods enable separation of the mixture under consideration into individual fractions of high purity indispensable for obtaining readable mass spectra. The purpose of this paper is to determine applicability of these methods in analysis of protein composition of urine samples.

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Quantitative amino acid and proteomic analysis: Very low excretion of polypeptides >750 Da in normal urine

Abstract: There is only very low excretion of polypeptides >750 Da in normal human urine. In Fanconi syndrome, excretion of unknown peptides of mass 2 to 5 kD, possibly relevant to the development of renal failure, is greatly increased.

Cited by 74 publications

References 23 publications

Reduced Renal Methylarginine Metabolism Protects against Progressive Kidney Damage

Reduced Renal Methylarginine Metabolism Protects against Progressive Kidney Damage

Current Issues in Measurement and Reporting of Urinary Albumin Excretion

Applicability of 2D gel electrophoresis and liquid chromatography in proteomic analysis of urine using mass spectrometry MALDI-TOF

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