As urine is less complex than plasma and accumulates substantial physiological and pathophysiological changes from both plasma and the urinary system, it is regarded as an ideal source of sample for biomarker studies.
Methods
To gain a better understanding of the biological functions of urinary proteins and their association with human diseases, we did an in-depth urinary proteome analysis, using urine samples from 24 healthy individuals, that was based on multiple separation strategies, including direct one-dimensional liquid chromatography–tandem mass spectrometry (1DLC/MS/MS), two-dimensional LC/MS/MS, and gel-eluted liquid fraction entrapment electrophoresis (GELFrEE) and liquid-phase isoelectric focusing (LP-IEF) followed by 2DLC/MS/MS.
Findings
We identified 6412 proteins, of which 2213 had not been reported in previous studies and 2571 were relatively and absolutely quantitated (spanning a magnitude of 106) with an intensity-based absolute quantification algorithm. Tissue-based analysis revealed that 514 tissue-enriched proteins from 27 tissues and 917 tissue-enhanced proteins from 32 tissues could be detected in urine, suggesting that the urinary proteome might have an important role in studying the physiological and pathological changes of the related organs, such as the brain and kidney. We selected potential urinary biomarkers that were previously identified for glioma and IgA nephropathy by filtering biomarkers from the publicly accessible Human Urine Proteome Database and verified them by Western blotting. We also observed that deeper separation allowed identification of more tissue-related and disease-specific proteins. All the detailed information—such as protein name, accession number, peptide sequence, sequence coverage, and biomarker application—could be obtained from the Human Urine Proteome Database, which provides information for prediction of urinary candidate biomarkers and modification of targeted proteomics workflow.
Interpretation
Urine might indicate the physiological and pathological changes of several organs and could therefore be used as a promising source of sample for biomarker discovery, especially for brain and kidney diseases.
Funding
This work was supported by the National Basic Research Program of China (2013CB530805 and 2014CBA02005), the Key Basic Research Program of the Ministry of Science and Technology of China (2013FY114100), the National Natural Science Foundation of China (31200614 and 31400669), and the Beijing Natural Science Foundation (5132028).
