Specific Urinary Proteins Research Articles

Measurement of urinary protein in children

Proteinuria is an early hallmark of kidney disease and a major risk factor for systemic cardiovascular diseases. There are several methods to measure proteinuria, such as the urine dipstick test, 24-hour urinary protein excretion method, and spot urine for the protein-to-creatinine ratio. The urine dipstick test is simple but inaccurate. The 24-hour urinary protein excretion method is the gold standard; however, it is cumbersome, especially in children. Spot urine for the protein-to-creatinine ratio is simple and accurate, but has limitations. Specific urinary protein such as albumin can be measured instead of the total protein content. Tests should be avoided in situations that cause transient proteinuria or false-positive results. It should be performed correctly, and its limitations should be recognized and interpreted accurately.

Computational Integration of Renal Histology and Urinary Proteomics using Neural Networks.

Histological image data and molecular profiles provide context into renal condition. Often, a biopsy is drawn to diagnose or monitor a suspected kidney problem. However, molecular profiles can go beyond a pathologist's ability to see and diagnose. Using AI, we computationally incorporated urinary proteomic profiles with microstructural morphology from renal biopsy to investigate new and existing molecular links to image phenotypes. We studied whole slide images of periodic acid-Schiff stained renal biopsies from 56 DN patients matched with 2,038 proteins measured from each patient's urine. Using Seurat, we identified differentially expressed proteins in patients that developed end-stage renal disease within 2 years of biopsy. Glomeruli, globally sclerotic glomeruli, and tubules were segmented from WSI using our previously published HAIL pipeline. For each glomerulus, 315 handcrafted digital image features were measured, and for tubules, 207 features. We trained fully connected networks to predict urinary protein measurements that were differentially expressed between patients who did/ did not progress to ESRD within 2 years of biopsy. The input to this network was either glomerular or tubular histomorphological features in biopsy. Trained network weights were used as a proxy to rank which morphological features correlated most highly with specific urinary proteins. We identified significant image feature-protein pairs by ranking network weights by magnitude. We also looked at which features on average were most significant in predicting proteins. For both glomeruli and tubules, RGB color values and variance in PAS+ areas (specifically basement membrane for tubules) were, on average, more predictive of molecular profiles than other features. There is a strong connection between molecular profile and image phenotype, which can be elucidated through computational methods. These discovered links can provide insight to disease pathways, and discover new factors contributing to incidence and progression.

Взаимосвязь содержания специфического протеина SERPINA1 в моче беременных с тяжестью преэклампсии и перинатальными исходами

Взаимосвязь содержания специфического протеина SERPINA1 в моче беременных с тяжестью преэклампсии и перинатальными исходами

Discovery of Novel Protein Biomarkers in Urine for Diagnosis of Urothelial Cancer Using iTRAQ Proteomics

Urothelial carcinoma (UC) is the ninth most prevalent malignancy worldwide. Noninvasive and efficient biomarkers with high accuracy are imperative for the surveillance and diagnosis of UC. CKD patients were enrolled as a control group in this study for the discovery of highly specific urinary protein markers of UC. An iTRAQ-labeled quantitative proteomic approach was used to discover novel potential markers. These markers were further validated with 501 samples by ELISA assay, and their diagnostic accuracies were compared to those of other reported UC markers. BRDT, CYBP, GARS, and HDGF were identified as novel urinary UC biomarkers with a high discrimination ability in a population comprising CKD and healthy subjects. The diagnostic values of the four novel UC markers were better than that of a panel of well-known or FDA-approved urinary protein markers CYFR21.1, Midkine, and NUMA1. Three of our discovered markers (BRDT, HDGF, GARS) and one well-known marker (CYFR21.1) were finally selected and combined as a marker panel having AUC values of 0.962 (95% CI, 0.94-0.98) and 0.860 (95% CI, 0.83-0.89) for the discrimination between UC and normal groups and UC and control (healthy + CKD) groups, respectively.

P0489URINARY PROTEOMIC MARKERS OF MEMBRANOUS NEPHROPATHY

Abstract Background and Aims Membranous nephropathy (MN) is considered one of the major conditions causing nephrotic-range proteinuria. Last years brought a huge progress in the field of non-invasive diagnostics, mainly due to the PLA2R antibody use in diagnostics and follow-up of primary MN. However, our understanding of MN is far from complete, especially that PLA2R-antibodies are found only in approximately 60-70% of all cases. Urinary proteomics is a promising tool that can help us to improve diagnostics, monitoring and understanding of MN. Our previous studies (Patent No. WO/2017/212463) suggested that specific urinary proteins can be used in the diagnostics of IgA nephropathy, lupus nephritis and ADPKD. The aim of this study was to evaluate with Mass Spectrometry (MS) patients diagnosed with MN in comparison to healthy controls. Method This study included patients with biopsy-proven MN (25) and healthy controls (7). Urine samples were obtained from a midstream of the second- or third-morning (SPOT) sample. The samples were processed up to 2 h after collection and stored at -80°C for further measurements with MS. The results were related to demographic data, standard laboratory tests and GFR estimated with use of Chronic Kidney Disease Epidemiology Collaboration (CKD-EPI) equation. Results The signal intensity of urine protein A, B and C (patent pending and the name of proteins disclosed until the date of ERA-EDTA Congress) was found to be significantly higher (p < 0.05) in MN patients compared to healthy controls (Fig. 1). Mass spectrometry, according to the specific amino-acids fragments of each tested protein, confirmed the differences between tested and control group. Additionally, statistically significant differences exist between patients with different types of glomerulonephritides (comparative MS in progress). Conclusion The signal intensity of urine proteins A, B, C is elevated in MN and according to our study, vary depending on types of nephropathies. This observation might suggests their differential roles in the pathophysiology of MN and its’ possible application as a non-invasive diagnostic and prognostic marker. Further studies are desired to establish the role of these urinary proteins.

P0768NOVEL URINARY PROTEIN BIOMARKERS FOR UROTHELIAL CANCER IN PATIENTS WITH CHRONIC KIDNEY DISEASE

Abstract Background and Aims Introduction: Chronic kidney disease (CKD) patients are known to have greater risk of urothelial cancer (UC) than general population. Urine is the most non-invasive examination and is directly linked to UC. Urinalysis and urine cytology are the major screening tools; however, these tests have low sensitivity or specificity in early disease. In this study, we used both normal and Non-UC CKD patients as controls to discover novel and more specific protein biomarkers in urine for early UC diagnosis. Aims To discover novel and more specific urinary protein biomarkers for screening of urothelial cancer (UC) in CKD patients. Method This is a multicenter prospective cohort study, carried out at 10 nationwide hospitals covering different regions of Taiwan from 2013-2018. Subjects were Taiwanese healthy adults (healthy controls, n=210), patients with CKD but without UC (disease controls, n=273) and patients with ongoing UC (n=211). For non-UC patients, urine and blood specimens were collected at the time of the structured interview. For UC patients, urine and blood samples were collected before operation. iTRAQ-LC-MS/MS approach was applied to compare protein expression among normal, CKD and UC cohorts. Results A total of 3838 proteins in urine were identified and 2497 proteins of them can be quantified. By filtering with fold changes, published papers and Human ATLAS, 27 candidates were selected as potential biomarkers and were validated with ELISA assay in a large sample size of urine specimens. Among the 27 candidates, we found BRDT, CYBP, GARS and HDGF were highly-expressed in the UC cohort when compared to normal and CKD cohorts (Figure 1). To increase the diagnostic value of UC, we combined ROC of our newly discovered 4 protein biomarkers. The AUC values of our newly discovered 4-biomarker panel were higher than the AUC values of the published biomarker panel, including the FDA-approved protein biomarkers (Figure 2). Conclusion We discover and validate a highly specific urinary protein biomarker panel (BRDT, GARS, HDGF and CYBP) to distinguish UC from CKD and healthy controls. It may be used for high throughput screening of subjects, such as CKD patients, who are at high risk to develop UC.

Proteinuria is Associated with Carotid Artery Atherosclerosis in Non-Albuminuric Type 2 Diabetes: A Cross-Sectional Study.

The association of specific urinary proteins other than albumin with cardiovascular (CV) outcomes in patients with type 2 diabetes (T2D) has been shown. In this respect, CV outcomes may differ in non-albuminuric T2D patients who were considered as a low risk group, according to the presence of proteinuria. We investigated the association between proteinuria and atherosclerosis assessed by carotid artery intima-media thickness (CIMT) in non-albuminuric T2D patients. 2047 T2D patients whose urine albumin-to-creatinine ratio was below 30 mg/g were recruited and classified into a non-proteinuria (NP, uPCR < 150 mg/g, n = 1865) group and a non-albuminuric proteinuria (NAP, uPCR ≥ 150 mg/g, n = 182) group. CIMT was compared between the two groups and logistic regression analysis was conducted to verify whether proteinuria could predict deteriorated CIMT status. In this cross-sectional study, mean CIMT of the NAP group were significantly thicker than those of the NP group (0.73 ± 0.16 vs. 0.70 ± 0.14, p = 0.016). The presence of proteinuria is associated with deteriorated CIMT after the adjustment for conventional risk factors (odds ratio, 2.342; 95% confidence interval, 1.082–5.070, p = 0.030) in regression analysis. We postulated that the measurement of urinary protein in conjunction with albumin might be helpful for predicting atherosclerosis, especially for non-albuminuric patients.

UBC®RapidTest for detection of carcinoma in situ for bladder cancer

UBC® Rapid Test is a test that detects fragments of cytokeratins 8 and 18 in urine. We present results of a multicentre study measuring UBC® Rapid Test in bladder cancer patients and healthy controls with focus on carcinoma in situ (CIS) and high-grade bladder cancer. From our study with N = 452 patients, we made a stratified sub-analysis for carcinoma in situ of the urinary bladder. Clinical urine samples were used from 87 patients with tumours of the urinary bladder (23 carcinoma in situ, 23 non-muscle-invasive low-grade tumours, 21 non-muscle-invasive high-grade tumours and 20 muscle-invasive high-grade tumours) and from 22 healthy controls. The cut-off value was defined at 10.0 µg/L. Urine samples were analysed by the UBC® Rapid Test point-of-care system (concile Omega 100 POC reader). Pathological levels of UBC Rapid Test in urine are higher in patients with bladder cancer in comparison to the control group (p < 0.001). Sensitivity was calculated at 86.9% for carcinoma in situ, 30.4% for non-muscle-invasive low-grade bladder cancer, 71.4% for nonmuscle-invasive high grade bladder cancer and 60% for muscle-invasive high-grade bladder cancer, and specificity was 90.9%. The area under the curve of the quantitative UBC® Rapid Test using the optimal threshold obtained by receiveroperated curve analysis was 0.75. Pathological values of UBC® Rapid Test in urine are higher in patients with high-grade bladder cancer in comparison to low-grade tumours and the healthy control group. UBC® Rapid Test has potential to be more sensitive and specific urinary protein biomarker for accurate detection of high-grade patients and could be added especially in the diagnostics for carcinoma in situ and non-muscle-invasive high-grade tumours of urinary bladder cancer.

Progress in urinary protein tumor markers

The development of proteomic technique has sparked new focus on novel protein markers for many diseases including cancer. Urine is one of the most useful biofluids for routine testing to provide an ideal resource for the discovery of novel biomarkers, with the advantage over tissue biopsy samples due to the ease and the less invasive nature of urinary collection. This review summarizes urinary protein tumor markers found in prostate cancer, bladder cancer, lung cancer and breast cancer that have shown potential for urogenital and non-urogenital cancer, so as to provide reference on high sensitive and high specific urinary protein tumor markers for early prevention, early diagnosis and treatment of cancer. Key words: Urinary protein; Tumor markers; Early diagnosis

Urine Protein Biomarker Candidates for Autism

Autism Spectrum Disorder (ASD) is increasingly common and treatment is most successful if instituted early in life. Urine biomarkers of autism could hasten diagnosis as urine is easy to obtain non-invasively. The purpose of this research was to compare urine samples from 8 ASD subjects and 8 age- and gender-matched controls. Samples were analyzed with isobaric Tags with Relative and Absolute Quantitation (iTRAQ™), a mass spectrometry method which enables identification and relative quantification of many proteins. We identified 231 proteins present in at least two control and two Autism subjects for statistical analysis. We ranked the proteins according to the P-values between Autism and control groups. The top five proteins were increased in ASD subjects compared to controls and included alpha 1-acid glycoprotein, prostaglandin-H2 D-isomerase, kininogen-1 isoform 2, leucine-rich alpha-2- glycoprotein 1 and immunoglobulin fragment Fab New lambda light chain. Two of the top 10 proteins have previously been related to autism, while six have previously been related to inflammation. We analyzed the 231 proteins with Ingenuity Pathway Analysis (IPA) to assess pathways involved and potential biomarkers. 104 of the 231 proteins were suggested by IPA as possible biomarkers in urine. The remaining 127 urinary proteins we identified are novel as they are not included as IPA urinary proteins. These research data fit with some current hypotheses regarding autism and suggest a relationship between ASD, inflammation and gastrointestinal disease. Specific urinary proteins are identified which could potentially serve as biomarkers for ASD.

Pursuing type 1 diabetes mellitus and related complications through urinary proteomics

Diabetes mellitus is a chronic metabolic disease with multiple complications, and its successful management requires early diagnosis, to allow timely interventions. Here, we have comprehensively analyzed the proteome changes in urine of type 1 diabetic subjects with and without complications such as retinopathy and nephropathy. gel electrophoresis combined to liquid chromatography-tandem mass spectrometry (GeLC-MS/MS) analysis of midstream urine highlighted the mechanisms involved in disease pathogenesis as, for instance wound healing and blood coagulation in all diabetics or altered ganglioside metabolism in retinopathy, and also some urinary proteins with potential diagnosis value. From these, gelsolin and antithrombin-III appear as promising diagnosis markers for type 1 diabetes mellitus (T1DM), whereas ephrin type-B receptor 4 and vitamin K-dependent protein Zseem to be promising markers for advanced T1DM disease state presenting retinopathy and nephropathy (T1DM-R+N). Data also suggest urinary ganglioside GM2 activator and beta-hexosaminidase subunit beta as potential urinary markers ofretinopathy in diabetics. Taken together, the present exploratory urinary proteomicanalysis might be seen as an important starting point for studies targeting specific urinary proteins aimed at the implementation of new biomarkers for the early detection of T1DM-related microvascular complications.

Clinical value of urinary kidney biomarkers for estimation of renal impairment in elderly Chinese with essential hypertension

The purpose of this work was to observe the excretion of specific types of urinary proteins and urinary enzymes in elderly essential hypertension patients, for early detection and targeted treatment of hypertensive nephropathy in the elderly. A total of 120 elderly essential hypertensive patients and 38 healthy elderly volunteers were involved. The urinary excretion rate of retinal-binding protein (RBP), transferrin (Tf), albumin (Alb), and urinary enzyme N-acetyl-beta-D-glucosaminidase (NAG) activity were determined. Patients were divided into two groups according to their creatinine clearance (Cockroft-Gault formula). There were 88 patients in group A, whose glomerular filtration rate (GFR) was >or=80 mL/min, and 32 patients in group B with a GFR <80 mL/min. Among the essential hypertensive patients, urinary excretion rates of RBP, Alb, Tf, and NAG were increased in both groups compared with the healthy controls. But the amount of urinary protein differed between group A and group B. The excretion rate of specific urinary protein and urinary enzyme had a positive relationship with the duration of course of hypertension. We believe that specific types of urinary proteins and urinary enzymes may be useful markers for early diagnosis of hypertensive nephropathy; they can also be regarded as a clinical indicator of the progression of hypertensive nephropathy, serving in the assessment of therapeutic effects.

Immunonephelometric quantification of specific urinary proteins versus a simple electrophoretic method for characterizing proteinuria

Objectives The quantification of urinary proteins presenting different molecular sizes is useful in characterizing a proteinuria. We assessed the performance of an electrophoretic system, the Hydragel Urine Profile, which allows firstly the identification of proteinuria and secondly a qualitative detection of monoclonal free light chains (FLC). Design and methods Initially, the proteinuria was characterized on 127 pathological urines by quantifying albumin, a1microglobulin, immunoglobulins G by immunonephelometric quantification technique and the results were compared with the protein pattern obtained by the electrophoretic method. Secondly, we assessed the sensitivity and specificity of this electrophoretic test for the detection and characterization of Bence Jones proteins. FLC were analyzed quantitatively by an immunonephelometric assay and qualitatively by the electrophoretic test in 150 urines. Results The agreement between the two methods was good with a percentage of homology for characterizing the proteinuria of 89%. For detecting a monoclonal FLC, the electrophoretic method demonstrated a lesser sensitivity but a higher specificity compared to the immunoassay. Conclusion The Urine Profile kit is a reliable assay that can be used as a screening test to differentiate the type of proteinuria.

Urine proteomics: the present and future of measuring urinary protein components in disease

For centuries, physicians have attempted to use the urine for noninvasive assessment of disease. Today, urinalysis, in particular the measurement of proteinuria, underpins the routine assessment of patients with renal disease. More sophisticated methods for assessing specific urinary protein losses have emerged; however, albumin is still the principal urinary protein measured. Changes in the pattern of urinary protein excretion are not necessarily restricted to nephrourological disease; for instance, the appearance of beta-human chorionic gonadotropin in the urine of pregnant women is the basis for all commercially available pregnancy kits. Similarly, microalbuminuria is a clinically important marker not only of early diabetic nephropathy but also of concomitant cardiovascular disease. With the emergence of newer technologies, in particular mass spectrometry, it has become possible to study urinary protein excretion in even more detail. A variety of techniques have been used both to characterize the normal complement of urinary proteins and also to identify proteins and peptides that may facilitate earlier detection of disease, improve assessment of prognosis and allow closer monitoring of response to therapy. Such proteomics-based approaches hold great promise as the basis for new diagnostic tests and as the means to better understand disease pathogenesis. In this review, we summarize the currently available methods for urinary protein analysis and describe the newer approaches being taken to identify urinary biomarkers.

Diagnostic performance of combined specific urinary proteins and urinary flow cytometry in urinary tract pathology

Urinalysis comprises three techniques: urinary flow cytometry, test strip analysis and determination of specific urinary proteins. We investigated the diagnostic possibilities of combining these methods for a cohort of patients with a variety of well-documented urological and nephrological pathology. Urinary samples from 407 in- and out-patients with nephrological or urological pathology were retrospectively included in our study. Test strip analysis (URISYS 2400), urinary flow cytometry (UF-100) and urinary protein analysis [albumin, total protein, alpha1-microglobulin (A1M), alpha2-macroglobulin (A2M)] were performed. In discriminating upper and lower urinary tract infections, A1M and A1M/log(white blood cells) can be used, whereas pathological casts only give poor discrimination. The ratio A2M/log(red blood cells; RBC) allows differentiation between cystitis and pyelonephritis, while glomerular diseases can be recognised by the log(RBCxurinary total protein). Combining A2M and urinary albumin allows the determination of acute prostatitis. Using combined parameters provided by various urinalysis techniques, ratios can be proposed with diagnostic value for classifying renal and urological conditions. Possible integration in computer-based knowledge systems may offer valuable information for clinicians dealing with these pathologies.

Apport du dosage pondéral par immunonéphélémétrie de différentes protéines urinaires pour l'interprétation des protéinuries

Detection and classification of proteinuria are frequently expected in clinical practice. Actually, these tests are realized by electrophoresis, which gives a panoramic view of urine proteins. However, this method remains semi-quantitative and its interpretation is investigator-dependant. Quantitative measurements of urine proteins have been developed but these techniques are not widely used despite their simplicity, specificity, sensitivity and rapidity. The aim of this article was to give a new approach of proteinuria interpretation by specific measurement of urine proteins followed by a software interpretation. Development of software to classify patterns of specific urinary protein markers improves the quality of follow-up care for patients and makes the interpretation easier.

Interpreting complex urinary patterns with mdi lablink: a statistical evaluation

The measurement of urinary marker proteins is not a generally accepted laboratory practice because the results are difficult to interpret. mdi-lablink is software for classifying patterns of specific urinary marker proteins. The interpretations are completely user definable thanks to a specific ‘pattern definition database’. Our interpretation set is based on Hofmann and Guder’s work in measuring and interpreting single urinary proteins. We include additional marker proteins in order to adapt Boesken’s SDS–PAGE classification. During the last 3 years, 1905 patterns were fully differentiated and identically interpreted. Firstly, the samples were classified into three patterns: normal (25.8%), predominantly glomerular (27.2%, selective, unselective, mixed, and with additional tubular proteins) and predominantly tubular (36.9%, complete/incomplete form, with additional glomerular proteins); 8.9% showed postrenal proteinuria. Secondly, glomerular selectivity measured by using urinary transferrin/IgG ratio alone correlates well with the established SI index (the ratio between IgG and transferrin clearances). Thirdly, the creatinine concentration substantiates the validity of the sample. The quality of the preanalytical phase can be improved through the ongoing education of the medical staff. Finally, measurement of urinary albumin and α-1-microglobulin is mandatory where kidney disease is suspected, has to be ruled out, or requires close monitoring, even when the total protein concentration is normal.

The role of α2u-globulin in ochratoxin A induced renal toxicity and tumors in F344 rats

The mycotoxin ochratoxin A (OTA) was shown to be a potent kidney carcinogen in rats demonstrating a marked sex difference in the response. Compared to female rats, male rats had a 10-fold higher incidence of kidney carcinomas. The objective of this study was to investigate whether this sex difference in tumor response is due to an exacerbation of effect resulting from the interaction of the male rat specific urinary protein α2u-globulin ( α2u) with OTA. Male and female rats were treated by oral gavage with OTA (1 mg/kg per day), d-limonene (dL; 1650 mg/kg per day) as a positive control or corn oil for 7 consecutive days. OTA induced severe renal lesions predominantly in the P 3 region of the proximal tubules. The lesions consisted of necrotic cells and cell exfoliations. No hyaline droplets were found in the P 2 segment following OTA treatment, whereas dL induced the expected accumulation of droplets. The results suggest that OTA induced kidney lesions are in all characteristic points different from the known α2u-nephropathy induced by dL. Based on these experiments the male rat specific protein α2u does not seem to be involved in the mechanism(s) leading to the high tumor incidence observed in OTA exposed male rats.

Effect of Plasma Replacement Therapy on Determinations of Urine Protein Concentration

Total protein concentration in urine is currently analyzed in our laboratory by a colorimetric method—Pyrogallol Red (PR)–molybdate complex (1)—adapted to a Cobas Integra analyzer (Cobas Integra Total Protein Urine/CSF 07.5723.3; Roche). We quantify specific urinary proteins by the following immunochemical methods: turbidimetry for urinary albumin (Cobas Integra) and nephelometry for IgG, α1-microglobulin, and light chains of immunoglobulins (BNAII; Behring). We recently observed discrepancies between results obtained for some urine samples, in that the total protein concentration was increased and did …

Use of Specific Urinary Proteins as Diagnostic Markers for Renal Disease

Use of Specific Urinary Proteins as Diagnostic Markers for Renal Disease