Identification Of Urinary Proteins Research Articles

Urinary Proteome of Newborn Calves-New Potential in Non-Invasive Neonatal Diagnostic.

Simple SummaryUrine is an underappreciated biological sample in the context of health monitoring of newborn calves. It can be collected noninvasively and used for diagnostic purposes, which is an attractive option in veterinary examination. The establishment of urinary protein maps of healthy newborn calves is important for diagnosing and monitoring the progression of various diseases. The aim of this study was to characterize the urinary proteome profile of healthy newborn calves. We applied 2-D gel electrophoresis combined with MALDI-TOF-TOF-MS/MS and 1-D gel electrophoresis coupled with LC-MS-MS analysis to separate and identify proteins present in the urine of newborn calves. Urinary proteomic analysis allowed us to detect proteins which are characteristic of embryonic and neonatal development and proteins involved in the cardiovascular and digestive system, and bone and kidney development. Moreover, we identified proteins that can be used to assess normal glomerular and tubular physiology. In calves, one of the main causes of mortality in the neonatal period is diarrhea, affecting fluid and electrolyte balance. We also identified proteins involved in the renal transport of water and electrolytes, which can be useful in the early diagnosis and prevention of these type of disorders. Urine is a biological diagnostic material suitable not only for the analysis of kidney and urinary tract functions but also the function of other tissues and organs. The urine proteome of adult mammals differs from the urine proteome of neonatal ones. The establishment of urinary protein maps of healthy newborn calves is important for diagnosing and monitoring the progression of various diseases. The experiment was carried out on a Polish-Friesian var. of Black-and-White male calves in the sixth day of postnatal life. The two proteomics approaches used for separation and identification of urinary proteins were: 2-DE with MALDI-TOF-TOF-MS/MS and 1-DE with LC-MS/MS. This resulted in the identification of 692 urinary proteins. The majority of them were classified as extracellular proteins (40.32%), as well as proteins involved in regulation of major cellular processes (31.07%). We have observed the presence of unique proteins associated with embryonic (ameloblastin, alpha-fetoprotein, Delta-like protein, embryo-specific fibronectin 1 transcript variant, Indian hedgehog homolog) and kidney development (angiotensin-converting enzyme, angiotensinogen, aquaporin-1, calbindin, glypican 3, nidogen 1, pro-cathepsin H). Additionally, proteins involved in the renal regulation of water and electrolyte balance (angiotensinogen, angiotensin-converting enzyme, aquaporin-1, ezrin, uromodulin) were detected. Presented in the current study 1-D and 2-D urinary proteomic maps are the basis for the identification and detection of prognostic biomarkers important for defining a calf’s health status.

Urinary Proteomics in Nephrotic Syndrome

Nephrotic syndrome is the commonest glomerular disease. Typical symptoms could be proteinuria, low serum albumin and oedema. The mechanism of proteinuria in nephrotic syndrome is a defective glomerular filtration barrier. Renal biopsy is the gold standard for diagnosis of nephrotic syndrome currently which is invasive and based on histopathological features, therefore it seems to be necessary to search for noninvasive biomarkers to be used as the complementary tests in the diagnostics and prognostics of glomerular diseases, particularly when renal biopsy is limited or contraindicated. While a big proportion of urinary proteins originate from kidney tissue and these tissue specific proteins excrete more in kidney injury, therefore the identification of urinary proteins can further our understanding of renal dysfunction and renal disease including nephrotic syndrome. The interest of scientist to urinary proteomics is also growing for biomarker discovery. This review focuses on some types of nephrotic syndrome and proteomic studies applying urine specimen which have been reported.

Mini-Slab SDS–PAGE and identification of urinary proteins

Mini-Slab SDS–PAGE and identification of urinary proteins

Proteomic analysis of normal human urinary proteins isolated by acetone precipitation or ultracentrifugation

Proteomic techniques have recently become available for large-scale protein analysis. The utility of these techniques in identification of urinary proteins is poorly defined. We constructed a proteome map of normal human urine as a reference protein database by using two differential fractionated techniques to isolate the proteins. Proteins were isolated from urine obtained from normal human volunteers by acetone precipitation or ultracentrifugation, separated by two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) and identified by matrix-assisted laser desorption ionization-time-of-flight (MALDI-TOF) mass spectrometry followed by peptide mass fingerprinting. A total of 67 protein forms of 47 unique proteins were identified, including transporters, adhesion molecules, complement, chaperones, receptors, enzymes, serpins, cell signaling proteins and matrix proteins. Acetone precipitated more acidic and hydrophilic proteins, whereas ultracentrifugation fractionated more basic, hydrophobic, and membrane proteins. Bioinformatic analysis predicted glycosylation to be the most common explanation for multiple forms of the same protein. Combining two differential isolation techniques magnified protein identification from human urine. Proteomic analysis of urinary proteins is a promising tool to study renal physiology and pathophysiology and to determine biomarkers of renal disease.

Separation and Identification of Urinary Proteins Following Digital Rectal Examination in Patients with Benign Prostatic Hypertrophy.

Separation and Identification of Urinary Proteins Following Digital Rectal Examination in Patients with Benign Prostatic Hypertrophy.

Separation and identification of urinary proteins and stone-matrix proteins by mini-slab sodium dodecyl sulfate-polyacrylamide gel electrophoresis.

Mini-slab electrophoresis on sodium dodecyl sulfate/polyacrylamide gel can be used to detect protein in unconcentrated urines containing more than 100 mg of protein per liter. Several urinary proteins--including albumin, transferrin, and the Tamm-Horsfall mucoprotein--precipitate, partly or totally, if urine samples are stored at 4 degrees C for one or two days. The present procedure permits identification of the albumin and transferrin in unconcentrated urines, thus avoiding their partial loss. Stone-matrix proteins in 10 mg or more of urinary calculi also could be detected. Albumin, transferrin, and Tamm-Horsfall mucoprotein accounted for greater than 40% of the total matrix protein in most of the urinary stones analyzed. The precipitation properties of these proteins and their high concentrations in stone matrix relative to those in urine suggest that they may play a role in stone formation and growth.

Analysis of proteinuria using a commercial system for automated electrophoresis and isoelectric focusing.

We describe an investigation of proteinuria using Pharmacia PhastSystemTM electrophoresis apparatus. The analysis of urinary proteins by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) of unconcentrated urine followed by silver staining took about 2 h and could clearly demonstrate tubular dysfunction or glomerular damage in urines with a negative or only trace-positive dip-stick test for protein. In addition, we show the identification of urinary proteins by immunoblotting from SDS-PAGE gels and the characterisation of Bence-Jones proteins by isoelectric focusing (IEF) and immunoblotting.