Urinary fetuin-A is a novel marker for diabetic nephropathy in type 2 diabetes identified by lectin microarray.
ABSTRACT: We analyzed the urine samples of patients with type 2 diabetes at various stages of diabetic nephropathy by lectin microarray to identify a biomarker to predict the progression of diabetic nephropathy. Japanese patients with type 2 diabetes at various stages of nephropathy were enrolled and we performed lectin microarray analyses (n = 17) and measured urinary excretion of fetuin-A (n = 85). The increased signals of urine samples were observed in Sia?2-6Gal/GalNAc-binding lectins (SNA, SSA, TJA-I) during the progression of diabetic nephropathy. We next isolated sialylated glycoproteins by using SSA-lectin affinity chromatography and identified fetuin-A by liquid chromatography-tandem mass spectrometer. Urinary excretion of fetuin-A significantly increased during the progression of albuminuria (A1, 0.40 ± 0.43; A2, 0.60 ± 0.53; A3 1.57 ± 1.13 ng/gCr; p = 7.29 × 10(-8)) and of GFR stages (G1, 0.39 ± 0.39; G2, 0.49 ± 0.45; G3, 1.25 ± 1.18; G4, 1.34 ± 0.80 ng/gCr; p = 3.89 × 10(-4)). Multivariate logistic regression analysis was employed to assess fetuin-A as a risk for diabetic nephropathy with microalbuminuria or GFR<60 mL/min. Fetuin-A is demonstrated as a risk factor for both microalbuminuria and reduction of GFR in diabetic nephropathy with the odds ratio of 4.721 (1.881-11.844) and 3.739 (1.785-7.841), respectively. Collectively, the glycan profiling analysis is useful method to identify the urine biomarkers and fetuin-A is a candidate to predict the progression of diabetic nephropathy.
Project description:Although increased urinary albumin excretion may increase the risk of adverse renal outcomes in patients with diabetes, it remains unclear whether microalbuminuria is associated with a higher incidence of macroalbuminuria in the absence of non-diabetic kidney events that frequently develop during the long-term course of type 2 diabetes. This historical cohort study included patients with type 2 diabetes, spot urine albumin:creatinine ratio (ACR) <300 mg/gCr and normal serum creatinine concentrations treated between August 1988 and April 2015. Patients with any evidence suggesting non-diabetic kidney diseases at baseline were excluded. Over a median follow-up of 50 months, 70 of the 1760 included patients developed macroalbuminuria. Twenty-one of these patients were diagnosed with non-diabetic renal events. The five-year cumulative incidence of macroalbuminuria in patients with ACRs of 0-7.5 mg/gCr, 7.5-30 mg/gCr, 30-150 mg/gCr, and 150-300 mg/gCr were 0%, 0.53%, 3.5%, and 36.0%, respectively, with significant differences between each pair of ACR categories. In type 2 diabetes, higher urinary ACR, even within a level of normoalbuminuria, was associated with a greater incidence of macroalbuminuria when non-diabetic renal events were excluded. These results conflict with findings suggesting that microalbuminuria is a poor indicator for the progression of diabetic nephropathy.
Project description:<h4>Background</h4>Improving the early detection of diabetic nephropathy remains a great challenge in disease management. Periostin is a marker of renal tubular injury and related to progressive kidney injury in animal models of chronic kidney disease. The clinical implications of urinary periostin activities in patients with type 2 diabetes have not been evaluated.<h4>Methods</h4>Urine samples were obtained from 30 healthy volunteers and 328 type 2 diabetic patients with normoalbuminuria (n=114), microalbuminuria (n=100) and macroalbuminuria (n=114). The excretion levels of urinary periostin were quantified with enzyme-linked immunosorbent assay. Immunohistochemical periostin expression was determined in kidney tissues from overt diabetic nephropathy.<h4>Results</h4>Increased periostin expression in glomeruli and tubular epithelium in diabetic renal pathology was observed. Urinary periostin levels were significantly elevated in the patients of the normoalbuminuria [3.06 (IQR: 1.12, 6.77) ng/mgCr], microalbuminuria [8.71 (IQR: 5.09, 19.29) ng/mgCr] and macroalbuminuria [13.58 (IQR: 3.99, 16.19) ng/mgCr] compared with healthy controls [1.15 (IQR: 0.60, 1.63) ng/mgCr] (P<0.01).Increased urine periostin level significantly correlated with aging, high albuminuria and decline of GFR. Urine periostin ELISA also demonstrated high performance for the diagnosis of established normoalbuminuric, microalbuminuric and macroalbuminuric type 2 diabetes (AUC 0.78 (95%CI, 0.71 to 0.86), 0.99 (95%CI, 0.98 to 1.00) and 0.95 (95%CI, 0.91 to 0.98), respectively).<h4>Conclusion</h4>The study indicates that increased urine periostin levels can be detected in patients with type 2 diabetes before the onset of significant albuminuria. Urinary periostin is an associated renal derangement in patients with established diabetic nephropathy and it may be used as an early marker of diabetic renal injury.
Project description:A number of proteomic and peptidomic analyses of urine from diabetic subjects have been published in the quest for a biomarker that predicts progression of nephropathy. Less attention has been paid to the relationships between urinary proteins and the underlying biological processes revealed by the analyses. In this review, we focus on the biological processes identified by studying urinary proteins and protein-protein interactions at each stage of diabetic nephropathy to provide an overview of the events underlying progression of kidney disease reflected in the urine. In uncomplicated diabetes, proteomic/peptidomic analyses indicate that early activation of fibrotic pathways in the kidney occurs before the onset of microalbuminuria. In incipient nephropathy, when albumin excretion rates are abnormal, proteomic/peptidomic analyses suggest that changes in glomerular permselectivity and tubular reabsorption account, at least in part, for the proteins and peptides that appear in the urine. Finally, overt nephropathy is characterized by proteins involved in wound healing, ongoing fibrosis, and inflammation. These findings suggest that there is a spectrum of biological processes in the diabetic kidney and that assessing protein networks may be more informative than individual markers with respect to the stage of disease and the risk of progression.
Project description:The poor performance of current tests for predicting the onset, progression and treatment response of diabetic nephropathy has engendered a search for more sensitive and specific urinary biomarkers. Our goal was to develop a new method for protein biomarker discovery in urine from these patients.We analyzed urine from normal subjects and patients with early and advanced nephropathy. Proteins were separated using a novel analysis process including immunodepletion of high-abundance proteins followed by two-stage LC fractionation of low-abundance proteins. The proteins in the fractions were sequenced using MS/MS.Immunodepletion of selected high-abundance proteins followed by two-stage LC produced approximately 700 fractions, each less complex and more amenable to analysis than the mixture and requiring minimal processing for MS identification. Comparison of fractions between normal and diabetic nephropathy subjects revealed several low-abundance proteins that reproducibly distinguished low glomerular filtration rate (GFR) from both high GFR diabetic and normal subjects, including uteroglobin, a protein previously associated with renal scarring.We developed a novel method to identify low-abundance urinary proteins that enables the discovery of potential biomarkers to improve the diagnosis and management of patients with diabetic nephropathy.
Project description:OBJECTIVE: To estimate the direct medical costs of hypertensive patients with type 2 diabetes by the level of proteinuria and to evaluate the differences between patients whose nephropathy did and did not progress. RESEARCH DESIGN AND METHODS: We identified 7,758 patients with diabetes and hypertension who had a urine albumin-to-creatinine ratio (UACR) during 2001-2003 and at least one follow-up UACR 3-5 years later. Patients were followed for up to 8 years for progression of nephropathy, which was defined by increasing levels of proteinuria: normoalbuminuria (UACR < 30 mg/g), microalbuminuria (30-299 mg/g), macroalbuminuria (?300 mg/g), and end-stage renal disease (dialysis or transplant). We calculated annualized inpatient, outpatient, pharmaceutical, and total medical costs incurred by patients after the baseline measure through 2008, comparing patients who did and did not progress to a higher nephropathy stage. We also compared pre- and postprogression costs among those whose nephropathy progressed. RESULTS: Patients with normoalbuminuria who progressed to microalbuminuria experienced an annualized change in baseline costs that was $396 higher (P < 0.001) than those who maintained normal albuminuria ($902 vs. $506). Among those with microalbuminuria, progression was significantly associated with a $747 difference (P < 0.001) in annualized change in outpatient costs compared with no progression ($1,056 vs. $309). Among patients who progressed, costs were 37% higher following progression from normoalbuminuria to microalbuminuria ($10,188 vs. $7,424; P < 0.001), and 41% higher following progression from microalbuminuria to macroalbuminuria ($12,371 vs. $8,753; P < 0.001). CONCLUSIONS: Progression of nephropathy was strongly associated with higher subsequent medical care costs in hypertensive patients with diabetes. Greater prevention efforts may reduce the substantial economic burden of diabetic nephropathy.
Project description:Cross-sectional studies suggest the association between diabetic nephropathy and the PPAR?2 Pro12Ala polymorphism of the peroxisome proliferator-activated receptor ?2 (PPAR?2). Prospective data are limited to microalbuminuria and no information on renal function is available to date. The present study evaluates the association between the Pro12Ala polymorphism of PPAR?2 and the progression of albuminuria and decay in glomerular filtration rate (GFR) in type 2 diabetes.We studied 256 patients with an average 5-year follow-up. Among others, urinary albumin excretion rate (UAER) was measured on spot sample, GFR was estimated with the CKD-EPI Equation.Baseline UAER and GFR were similar for carriers or non-carriers of the polymorphism. At follow-up no significant changes from baseline were observed for UAER or eGFR in carriers of the Pro12Ala polymorphism whereas a significant increase in UAER [17 (11.3-37.9) versus 24.5 (13.8-49.9) ?g/mg, p?<?0.006)] and a significant reduction in the eGFR (82.8?±?14.5 versus 80.3?±?17.3 ml/min/1.73, m(2) p?=?0.02), were observed in non carriers of the Pro12Ala polymorphism. Progression of nephropathy - defined according to a combined end point of UAER and eGFR- i.e. doubling of baseline UAER to at least 100 ?g/mg, or new onset microalbuminuria, or progression from micro to macroalbuminuria, or 25% reduction of eGFR, or annualized eGFR decline >3 ml/min/year - was significantly less frequent in Ala carriers than non carriers (11.4% vs 35.8%; p?<?0.01); HR adjusted for baseline age, AER, eGFR, HbA1c, diabetes duration and blood pressure was 0.32 (0.12-0.80).This study found that among patients with type 2 diabetes, the PPAR?2 Pro12Ala polymorphism is protective against progression of nephropathy and decay of renal function independent of major confounders.
Project description:<i>Aims</i>. Dipeptidyl-peptidase IV inhibitors (DPP-4i) are among the most popular oral antidiabetic agents. However, the effects of DPP-4i on diabetic nephropathy are not well-established. The aim of this study was to determine the renoprotective effects of DPP-4i, using albuminuria and glomerular filtration rate (GFR) as indicators, in type 2 diabetes mellitus (T2DM) patients. <i>Methods</i>. This retrospective observational cohort study used the clinical database of a tertiary hospital. The changes of urine albumin/creatinine ratio (UACR), estimated GFR (eGFR), and metabolic parameters after treatment were compared with the changes of those parameters before treatment using paired Student's <i>t</i>-test. <i>Results</i>. The mean UACR in the entire study population decreased to approximately 45?mg/g 1 year after DPP-4i treatment, while it was increased approximately 39?mg/g 1 year before DPP-4i treatment (<i>p</i> < 0.05). Patients with macroalbuminuria showed a significant reduction in albumin levels after DPP-4i treatment (<i>p</i> < 0.05); however, patients with microalbuminuria and normoalbuminuria did not show improvements in albuminuria levels after treatment. Although eGFR was not changed 1 year after DPP-4i treatment, reductions in eGFR were slowed in patients with microalbuminuria and reversed in the macroalbuminuria or normoalbuminuria groups, 4 years after treatment. <i>Conclusions</i>. Administration of DPP-4i reduces urine albumin excretion and mitigates reduction of eGFR in T2DM patients.
Project description:Netrin-1 was recently identified as an early diagnostic biomarker of chronic kidney disease (CKD) in an experimental animal model. However, its usefulness for early diagnosis of CKD in humans is unknown. The current study evaluated whether netrin-1 is increased in urine from human diabetic patients.Spot urine samples from healthy volunteers, diabetes without microalbuminuria, diabetes with microalbuminuria and diabetes with macroalbuminuria were collected after receiving consent. Netrin-1 in urine was quantified by enzyme-linked immunosorbent assay and the data analyzed to determine whether urinary netrin-1 significantly correlates with disease progression.Urinary netrin-1 levels were significantly increased in normoalbuminuric diabetic patients compared to healthy controls, and still further elevated in patients with microalbuminuria and overt nephropathy. Urinary netrin-1 was significantly associated with albuminuria and estimated glomerular filtration rate, independently of age and sex.Netrin-1 is detectable in urine from diabetic patients and may serve as a useful early diagnostic biomarker predicting the development of CKD in diabetes.
Project description:<h4>Background</h4>In patients with Type 1 Diabetes (T1D) who develop microalbuminuria, progressive decline in glomerular filtration rate (GFR) may be initiated by leakage into the urine of toxic proteins (txUPs). This study tested this hypothesis.<h4>Methods</h4>After archiving baseline urine, we followed T1D patients with microalbuminuria for 8-12 years to distinguish those in whom GFR declined (Decliners) and those in whom it remained stable (Non-decliners). Human proximal tubular cells (HK-2 cells) were grown in serum-free medium enriched with pooled urines from Decliners or Non-decliners. We determined genome-wide expression profiles in extracted mRNA.<h4>Results</h4>The two pooled urines induced differential expression of 312 genes. In terms of gene ontology, molecular functions of the 119 up-regulated genes were enriched for protein binding and peptidase inhibitor activities. Their biologic processes were enriched for defense response, responses to other organisms, regulation of cellular processes, or response to stress or stimulus, and programmed cell death. The 195 down-regulated genes were disproportionately represented in molecular functions of cation binding, hydrolase activity, and DNA binding. They were disproportionately represented in biological processes for regulation of metabolic processes, nucleic acid metabolic processes, cellular response to stress and macromolecule biosynthesis. The set of up-regulated genes in HK-2 cells overlaps significantly with sets of over-expressed genes in tubular and interstitial compartments of kidney biopsies from patients with advanced DN (33 genes in one study and 25 in the other compared with 10.3 expected by chance, p<10(-9) and p<10(-4), respectively). The overlap included genes encoding chemokines and cytokines. Overlap of down-regulated genes was no more than expected by chance.<h4>Conclusions</h4>Molecular processes in tubules and interstitium seen in advanced diabetic nephropathy can be induced in vitro by exposure to urine from patients with minimal microalbuminuria who subsequently developed progressive renal function decline, presumably due to putative txUPs.
Project description:Microalbuminuria provides the earliest clinical marker of diabetic nephropathy among patients with Type 1 diabetes, yet it lacks sensitivity and specificity for early histological manifestations of disease. In recent years microRNAs have emerged as potential mediators in the pathogenesis of diabetes complications, suggesting a possible role in the diagnosis of early stage disease. We used quantiative polymerase chain reaction (qPCR) to evaluate the expression profile of 723 unique microRNAs in the normoalbuminuric urine of patients who did not develop nephropathy (n = 10) relative to patients who subsequently developed microalbuminuria (n = 17). Eighteen microRNAs were strongly associated with the subsequent development of microalbuminuria, while 15 microRNAs exhibited gender-related differences in expression. The predicted targets of these microRNAs map to biological pathways known to be involved in the pathogenesis and progression of diabetic renal disease. A microRNA signature (miR-105-3p, miR-1972, miR-28-3p, miR-30b-3p, miR-363-3p, miR-424-5p, miR-486-5p, miR-495, miR-548o-3p and for women miR-192-5p, miR-720) achieved high internal validity (cross-validated misclassification rate of 11.1%) for the future development of microalbuminuria in this dataset. Weighting microRNA measurements by their number of kidney-relevant targets improved the prognostic performance of the miRNA signature (cross-validated misclassification rate of 7.4%). Future studies are needed to corroborate these early observations in larger cohorts.