Poor histological lesions in IgA nephropathy may be reflected in blood and urine peptide profiling.

BACKGROUND: IgA nephropathy (IgAN) is the most common primary glomerulonephritis worldwide, leading to renal failure in 15% to 40% of cases. IgAN is diagnosed by renal biopsy, an invasive method that is not risk-free. We used blood and urine peptide profiles as a noninvasive method of linking IgAN-associated changes with histological lesions by Oxford classification. METHODS: We prospectively studied 19 patients with biopsy-proven IgAN and 14 healthy subjects from 2006 to 2009, excluding subjects with crescentic glomerulonephritis and collecting clinical and biochemical data at the time of diagnosis and during follow-up (24 months). Histological lesions were evaluated by Oxford classification. Proteomic analysis was performed by combining magnetic bead (MB) technology and mass spectrometry (MALDI-TOF MS) to obtain peptide profiles. Doubling of serum creatinine was considered a variable of poor renal prognosis. RESULTS: We identified 55 peptides-13 in serum, 26 in plasma, and 16 in urine-that differentiated IgAN patients from healthy subjects. A significant association was noted between serum/plasma and urine peptides and histological findings-ie, tubulointerstitial damage, segmental glomerulosclerosis, and endocapillary injury. CONCLUSIONS: In patients with IgAN, the use of noninvasive approaches, such as blood and urine proteomics, can provide valuable information beyond that of standard diagnostic techniques, allowing us to identify blood and urine peptide profiles that are associated with poor histological lesions in IgAN patients.

Uromodulin and α(1)-antitrypsin urinary peptide analysis to differentiate glomerular kidney diseases.

BACKGROUND/AIMS: Glomerular kidney disease (GKD) is suspected in patients based on proteinuria, but its diagnosis relies primarily on renal biopsy. We used urine peptide profiling as a noninvasive means to link GKD-associated changes to each glomerular entity. METHODS: Urinary peptide profiles of 60 biopsy-proven glomerular patients and 14 controls were analyzed by combining magnetic bead peptide enrichment, MALDI-TOF MS analysis, and ClinProTools v2.0 to select differential peptides. Tentative identification of the differential peptides was carried out by HPLC-MS/MS. RESULTS: The HPLC-MS/MS results suggest that uromodulin (UMOD; m/z: 1682, 1898 and 1913) and α(1)-antitrypsin (A1AT; m/z: 1945, 2392 and 2505) are differentially expressed urinary peptides that distinguish between GKD patients and healthy subjects. Low UMOD and high A1AT peptide abundance was observed in 80-92% of patients with GKD. Proliferative forms of GKD were distinguished from nonproliferative forms, based on a combination of UMOD and A1AT peptides. Nonproliferative forms correlated with higher A1AT peptide levels - focal segmental glomerulosclerosis was linked more closely to high levels of the m/z 1945 peptide than minimal change disease. CONCLUSION: We describe a workflow - urinary peptide profiling coupled with histological findings - that can be used to distinguish GKD accurately and noninvasively, particularly its nonproliferative forms.

Messenger RNA expression of B7-1 and NPHS1 in urinary sediment could be useful to differentiate between minimal-change disease and focal segmental glomerulosclerosis in adult patients.

BACKGROUND: Podocyte proteins are involved in the pathogenesis of glomerular kidney disease (GKD). However, there is little information on messenger RNA (mRNA) expression patterns of B7-1 and NPHS1 in urinary sediment of patients with GKD. The objective of this study was to analyse the gene expression of B7-1 in urinary sediment and correlate it with the expression of podocyte-specific genes in patients with GKD. METHODS: Adult patients with proliferative and non-proliferative GKD, proteinuria and stable renal function, were included. A group of healthy subjects was used to determine normal levels of urinary markers and to obtain reference RNA. Biochemical, clinical and experimental procedures included measurement of creatinine level and total urinary protein, renal biopsy, identification of urinary podocytes, gene expression analysis of B7-1, NPHS1, NPHS2 and SyNPO genes and urinary B7-1 protein analysis by enzyme-linked immunosorbent assay. RESULTS: Between June 2006 and November 2009, 69 patients with GKD (median age: 46 ± 15 years, 64% men) and 14 healthy subjects (median age: 34 ± 12 years, 43% men) were included. In both groups, urinary mRNA levels of B7-1 and NPHS1 were significantly higher in patients with GKD compared to healthy subjects (P = 0.050 and P = 0.008, respectively). Regarding GKD subtypes, patients with focal segmental glomerulosclerosis (FSGS), but not patients with minimal change disease (MCD), had a significantly higher mRNA expression of B7-1 and NPHS1 than healthy subjects (P = 0.012 and P = 0.030, respectively). Patients with MCD had a significantly lower NPHS1 mRNA expression than patients with FSGS (P = 0.012). The B7-1:NPHS1 urinary mRNA ratio was significantly higher in patients with MCD compared with patients with FSGS (P = 0.027). CONCLUSION: mRNA expression analysis of B7-1 and NPHS1 in urinary sediment may be useful to differentiate between different histologic subtypes of GKD, particularly between MCD and FSGS.

Proteomic approach to the study of statin pleiotropy in kidney transplant patients.

BACKGROUND/AIMS: Statins are prescribed in kidney transplant recipients in order to manage dyslipidemia, a common complication in these patients. The efficacy of statins in reducing cholesterol levels has been accompanied by pleiotropic effects. Fifty-four kidney transplant patients were included in the present study, the objective of which was to ascertain the effect of 12 weeks of atorvastatin therapy (10 mg/day) on the patients' lipid profile, renal function, markers of inflammation and plasma peptide profile. METHODS: Biochemical variables were determined with a routine clinical laboratory analyzer, and the proteomic approach was based on magnetic particle-assisted sample processing coupled to mass spectrometry readout. RESULTS: Atorvastatin therapy improved the lipid profile of patients and caused significant changes in their plasma peptide profile; peptides with m/z 1063 and 1898 decreased after treatment and were identified as fragments derived from molecules involved in vascular inflammation, i.e. high-molecular-weight kininogen and complement factor C4, respectively. CONCLUSION: These findings may contribute to the growing body of evidence of the anti-inflammatory actions attributed to statins, by which these drugs could improve these patients' clinical status.

Increased Urinary Erythropoietin Excretion in Severe Sleep Apnea-Hipoapnea Syndrome: The Effect of CPAP. / El síndrome de apneas-hipoapneas del sueño (SAHS) grave incrementa la excreción urinaria de eritropoyetina. Efecto del tratamiento con CPAP.

INTRODUCTION: Tissue hypoxia stimulates the production of erythropoietin (EPO), the main effect of which is, in turn, to stimulate erythropoiesis. Sleep apnea-hypopnea syndrome (SAHS) is an entity characterized by repeated episodes of hypoxemia during sleep. OBJECTIVE: To analyze whether hypoxemia stimulated increased urinary excretion of EPO, and if so, to evaluate if treatment with continuous positive airway pressure (CPAP) can inhibit this phenomenon. METHODS: We studied 25 subjects with suspected SAHS who underwent a polysomnography study (PSG). EPO levels in first morning urine (uEPO) and blood creatinine and hemoglobin were determined in all patients. Patients with severe SAHS repeated the same determinations after CPAP treatment. RESULTS: Twelve subjects were diagnosed with severe SAHS (mean ± SD, AHI 53.1 ± 22.7). Creatinine and hemoglobin levels were normal in all subjects. uEPO was 4 times higher in the SAHS group than in the control group (1.32 ± 0.83 vs. 0.32 ± 0.35 UI/l, p <.002). CPAP treatment reduced uEPO to 0.61 ± 0.9 UI/l (p <.02), levels close to those observed in healthy subjects. No dose-response relationship was observed between severity of PSG changes and uEPO values. CONCLUSIONS: Patients with severe SAHS show increased uEPO excretion, but this normalizes after treatment with CPAP.

Urinary peptide profiling to differentiate between minimal change disease and focal segmental glomerulosclerosis.

BACKGROUND: Minimal change disease (MCD) and primary focal segmental glomerulosclerosis (FSGS) are the main causes of primary idiopathic nephrotic syndrome in children and adults, with diagnosis being essential for the appropriate choice of therapy and requiring renal biopsy. However, the presence of only normal glomeruli on renal biopsy of FSGS patients may lead to the misclassification of these patients as having MCD. The aim of this study was to (i) compare the peptide profile of MCD and FSGS patients with that of a group of healthy subjects, (ii) generate and validate a class prediction model to classify MCD and FSGS patients and (ii) identify candidate biomarkers of these glomerular entities by analysis of the urinary peptidome. METHODS: The urinary peptide profile was analyzed by magnetic bead-based technology combined with MALDI-TOF mass spectrometry in 44 patients diagnosed of MCD (n = 22) and FSGS (n = 22). The resulting spectra were compiled and analyzed using ClinProTools software. RESULTS: A class prediction model was developed to differentiate MCD and FSGS patients. The validation of this model correctly classified 81.8% (9/11) of MCD patients and 72.7% (8/11) of FSGS patients. Moreover, the signal with m/z 1913.60, identified as a fragment of uromodulin, and the signal with m/z 2392.54, identified as a fragment of alpha-1-antitrypsin, showed higher and lower peak areas, respectively, in FSGS patients compared with MCD patients. CONCLUSIONS: The simple, non-invasive technique described in the present study may be a useful tool to help clinicians by confirming diagnoses achieved by renal biopsy, thereby reducing misdiagnoses and avoiding the implementation of inappropriate therapies.

Magnetic bead-based proteomic technology to study paricalcitol effect in kidney transplant recipients.

Secondary hyperparathyroidism is a common complication in patients with chronic kidney disease and frequently persists after kidney transplantation. Paricalcitol, a selective vitamin D receptor activator, is indicated in the management of this disorder and recent evidences have suggested that this drug has other beneficial effects. Aiming to elucidate these effects, our study included 52 stable kidney transplant recipients randomized 2:1 to treatment with paricalcitol or to no treatment. Bone mineral parameters, kidney function and inflammatory status were assessed at baseline, at 3 and at 12 months. Moreover, a proteomic approach, based on magnetic beads technology coupled to MALDI-TOF mass spectrometry readout, was used to determine changes in patients' plasma peptidome. Patients treated with paricalcitol showed a significant decrease in parathyroid hormone and alkaline phosphatase levels, and an increase of bone mineral density and glomerular filtration rate. The proteomic analysis revealed a decrease in bradykinin after paricalcitol treatment, whereas 2 peptides identified as fragments of the complement factor C4 decreased only in those patients not treated with paricalcitol. These findings suggest that paricalcitol may offer additional benefits due to immunomodulatory effects via the kallikrein-kinin and complement systems.