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Use of Urinary Proteomics for the Discovery of Biomarkers of Renal Disease
overflow or post-renal type. When the glomerular filtration barrier is Glomerular Disease
damaged, glomerular proteinuria develops, characterised by proteins Thongboonkerd et al. studied several glomerular diseases (diabetic
the size of albumin (69kDa) or larger in the urine. Impaired proximal nephropathy, focal segmental glomerulosclerosis and lupus nephritis)
tubule reabsorption of proteins that are normally filtered by the using 2D-PAGE.
17
A total of 25 proteins were expressed differentially
glomeruli causes tubular proteinuria. Overflow proteinuria occurs among the groups. In normal urine, the spot intensities for albumin,
when an increased amount of plasma protein leads to filtration transferrin and α1-antitrypsin were significantly reduced, whereas
through the normal glomeruli at a rate that is beyond the capacity of those of kininogen and phorbolin-3 were greater compared with the
the tubules to reabsorb the proteins. Finally, post-renal proteinuria can urine from patients with glomerular disease.
result from inflammation anywhere in the urinary tract (i.e. renal
pelvis, ureter, bladder, prostate or urethra). Haubitz et al. studied 45 patients with IgAN and found a specific pattern
of polypeptide excretion.
18
Sensitivity and specificity were 100 and
Glomerular proteinuria is seen in primary glomerular diseases 90%, respectively, compared with healthy individuals, and 77 and 100%,
(i.e. glomerulonephritis or immunoglobulin A nephropathy [IgAN]) and respectively, compared with patients with membranous nephropathy.
in systemic disorders that cause glomerular damage (e.g. diabetes Treatment of the patients was associated with changes in the pattern,
mellitus, lupus erythematosus or infectious diseases). Tubular possibly indicating a therapeutic effect. In another study, 13 patients with
proteinuria appears in interstitial nephritis, acute tubular necrosis and IgAN and 12 normal controls were analysed using 2D-PAGE separation
renal tubular acidosis. Overflow proteinuria occurs in disorders such as and MALDI-TOF identification.
19
A total of 84 differentially expressed spots
Bence–Jones proteinuria and myoglobinuria. representing 59 proteins were identified. Further studies are needed to
Normal Urine Proteome
Thongboonkerd et al. used 2D-PAGE to separate urine proteins previously
A non-invasive biomarker of
precipitated by acetone or ultracentrifuged.
10
They defined a human
rejection would allow not only
urinary proteome map consisting of 67 proteins and their isoforms. Since
an early diagnosis, but also
then, other proteomic maps for human urine obtained with 2D-PAGE
have been developed, with the identification of 113 proteins and 150 frequent monitoring of
proteins.
12
Schaub et al. used SELDI-TOF MS to obtain a profile of human
immunosuppressive therapy.
urinary proteins, but failed to identify any protein.
13
Pisitkun et al. studied
the proteome of urinary exosomes (small vesicles that originate as internal
vesicles in multivesicular bodies and that are secreted from the cell when determine the potential pathogenic role of the proteins and their
the outer membrane of the multivesicular body fuses with the plasma usefulness in the diagnosis and monitoring of IgAN. As noted by Fliser et
membrane) using a combination of ultracentrifugation followed by al., many of the biomarkers identified in the urine of patients with renal
1D electrophoresis and nanospray LC-MS/MS, and identified 295 protein diseases are proteolytic fragments of larger proteins.
20
Apparently, specific
species.
14
Seventy-three of the identified proteins participate in membrane proteases in the urine may cleave these excreted proteins.
trafficking and 21 are associated with kidney disease or hypertension.
Acute Renal Allograft Rejection
Sun et al. precipitated urine proteins with acetone and used three Acute renal allograft rejection is currently diagnosed by renal biopsy. A
approaches to separate and identify the proteins: 1DE plus 1D non-invasive biomarker of rejection would allow not only an early
LC-MS/MS, direct 1D LC-MS/MS and 2D LC-MS/MS.
15
A total of 226 diagnosis, but also frequent monitoring of immunosuppressive
urine proteins were identified. One hundred and seventy-one proteins therapy. In recent years, proteomics has been applied in the search for
were identified using a proteomic approach for the first time, including biological markers of acute allograft rejection. To this end, the main
four male-specific proteins. Twelve low-molecular-mass proteins were technological proteomic approach has been SELDI-TOF.
21–24
The results
identified. Most urinary proteins had a molecular mass between 30 obtained by these authors reveal completely different biomarkers for
and 60kDa, and a pI between four and 10. The apparent molecular the same disease; this was attributed to the use of differential chip
masses of many proteins were different from the theoretical ones, surfaces and analyses of the data collected in these studies. Currently,
indicating their post-translational modification and degradation. proteomic studies in urine have not yielded biomarkers useful for the
Adachi et al. have recently identified 1,543 proteins in the urine of early detection of acute allograft rejection, although this field is
healthy people.
16
Surprisingly, nearly half of the annotated proteins currently attracting the attention of many researchers and it is hoped
were membrane proteins. Plasma membrane proteins are probably that in the near future these studies will yield useful results.
present in urine due to secretion in exosomes. To date, this is the most
complete protein list for normal human urine. The studies carried out Urological Cancers
so far have provided a broad set of human urinary proteins that may There are currently no routinely used circulating tumour markers for renal
serve as a template for comparisons with urinary protein maps in cancer. In urological cancers, urine represents a particularly useful fluid in
diseases in the search for biomarkers. which to examine tumour markers. Urological malignancies, including both
bladder cancer and renal cell carcinoma (RCC), have been examined using
The Urine Proteome in Renal Diseases urinary proteomic methods. Most bladder cancers are transitional cell
Proteomic analyses of urine have been applied to the search for carcinomas (TCC), with squamous cell carcinomas (SCC) being much less
biomarkers of renal diseases, including glomerular disease, acute renal common. Several studies have searched for urinary protein biomarkers of
allograft rejection, urological cancers and urolithiasis. renal cancer.
25–29
Among the proteins detected were psoriasin, the major
EUROPEAN RENAL DISEASE 2007 9
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